RESUMO
Hepatocyte nuclear factor-4alpha (HNF4alpha) exists in multiple isoforms that are generated by alternative promoter (P1 and P2) usage and splicing. Here we establish monoclonal antibodies (mAbs) for detecting P1 and P2 promoter-driven HNF4alpha, and evaluate their expression in normal adult human tissues and surgically resected carcinomas of different origins. Using immunohistochemical analysis, we demonstrate that, while P1 promoter-driven HNF4alpha is expressed in hepatocytes, small intestine, colon, kidney and epididymis, P2 promoter-driven HNF4alpha is expressed in bile duct, pancreas, stomach, small intestine, colon and epididymis. Altered expression patterns of P1 and P2 promoter-driven HNF4alpha were observed in gastric, hepatocellular and colorectal carcinomas. HNF4alpha was expressed in lung metastases from renal cell, hepatocellular and colorectal carcinoma but was not observed in lung tumours. The P1 and P2 promoter-driven HNF4alpha expression pattern of tumour metastases correlated with the primary site of origin. P1 promoter-driven HNF4alpha was also found in intestinal metaplasia of the stomach. These data provide evidence for the tissue distribution of P1 and P2 promoter-driven HNF4alpha at the protein level and suggest that HNF4alpha may be a novel diagnostic marker for metastases of unknown primary. We propose that the dysregulation of alternative promoter usage of HNF4alpha is associated with the pathogenesis of certain cancers.
Assuntos
Biomarcadores Tumorais/metabolismo , Transformação Celular Neoplásica/genética , Fator 4 Nuclear de Hepatócito/metabolismo , Neoplasias/metabolismo , Regiões Promotoras Genéticas , Animais , Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/imunologia , Feminino , Regulação Neoplásica da Expressão Gênica , Fator 4 Nuclear de Hepatócito/genética , Fator 4 Nuclear de Hepatócito/imunologia , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Masculino , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/imunologia , Proteínas de Neoplasias/metabolismo , Neoplasias/genética , Lesões Pré-Cancerosas/metabolismo , RNA Mensageiro/genética , RNA Neoplásico/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Neoplasias Gástricas/metabolismo , Distribuição Tecidual , Células Tumorais CultivadasRESUMO
Myosin is a functional protein associated with cellular movement, cell division, muscle contraction and other functions. Members of the myosin super-family are distinguished from the myosin heavy chains that play crucial roles in cellular processes. Although there are many studies of myosin heavy chains in this family, there are fewer on non-muscle myosin heavy chains than of muscle myosin heavy chains. Myosin is classified as type I (myosin I) or type II (myosin II). Myosin I, called unconventional myosin or mini-myosin, has one head, while myosin II, called conventional myosin, has two heads. We transfected myosin heavy polypeptide 9 (MYH9) into HeLa cells as a fusion protein with enhanced green fluorescent protein (EGFP) and analyzed the localization and distribution of MYH9 in parallel with those of actin and tubulin. The results indicate that MYH9 colocalizes with actin stress fibers. Since it has recently been shown by genetic analysis that autosomal dominant giant platelet syndromes are MYH9-related disorders, our development of transfected EGFP-MYH9 might be useful for predicting the associations between the function of actin polymerization, the MYH9 motor domain, and these disorders.
Assuntos
Células HeLa/metabolismo , Proteínas Motores Moleculares/metabolismo , Músculo Esquelético/metabolismo , Cadeias Pesadas de Miosina/metabolismo , Actinas/metabolismo , Técnica Indireta de Fluorescência para Anticorpo , Proteínas de Fluorescência Verde , Humanos , Indicadores e Reagentes/metabolismo , Proteínas Luminescentes/genética , Proteínas Luminescentes/metabolismo , Microscopia de Fluorescência , Proteínas Motores Moleculares/genética , Cadeias Pesadas de Miosina/genética , Transfecção , Tubulina (Proteína)/metabolismoRESUMO
The inhibitory effect of paralytic shellfish poison and tetrodotoxin on nerves from toxic and nontoxic crabs was examined. The toxins at concentrations of 10(-3) - 10(-4) M partially or completely inhibited the action potential of nerves isolated from the legs of toxic crab species (Zosimus aeneus, Atergatis floridus and Platypodia granulosa), but had no effect at 10(-6) M, the concentration at which the action potential of nerves from a nontoxic crab (Plagusia dentipes) was inhibited completely. A xanthid crab Daira perlata was intermediate in respect to the resistance to toxins. These results agree with the previous results obtained by i.p. administration of both toxins into those crabs.
Assuntos
Braquiúros/efeitos dos fármacos , Sistema Nervoso/efeitos dos fármacos , Neurotoxinas , Saxitoxina , Tetrodotoxina , Potenciais de Ação/efeitos dos fármacos , Animais , Técnicas In Vitro , Frutos do MarAssuntos
Alginatos/farmacologia , Fibrinólise/efeitos dos fármacos , Animais , Masculino , Ratos , Ratos EndogâmicosRESUMO
Specimens of a xanthid crab Atergatis floridus were collected from two adjacent areas in Kabira Bay, Ishigaki Island, Okinawa and compared with respect to toxicity and toxin composition. 'Reef specimens', which were collected from the reefs of Kabira Bay, showed an average toxicity score of 380 MU/g as paralytic shellfish poison, and a toxin composition consisting mainly of saxitoxin and a neosaxitoxin-related substance. On the other hand, 'Kojima specimens', which were collected from a small island in the bay, showed an average toxicity of 38 MU/g as tetrodotoxin, and a toxin composition consisting mainly of tetrodotoxin and related substance(s).
Assuntos
Braquiúros/análise , Toxinas Marinhas/isolamento & purificação , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia em Camada Fina , Eletroforese em Acetato de Celulose , Feminino , Japão , MasculinoRESUMO
Toxins were extracted from the xanthid crab Atergatis floridus inhabiting Ishigaki Island, Okinawa, and subjected to several types of chromatography, resulting in separation into gonyautoxin and saxitoxin fractions. Thin-layer chromatographic and electrophoretic analyses showed that the gonyautoxin fraction was composed of gonyautoxins 1-4, along with some unknown compounds. Gas chromatography-mass spectrometry demonstrated that the gonyautoxin fraction gave rise to the C9-base when alkali-hydrolyzed, indicating that this fraction contained a tetrodotoxin-like compound possessing the quinazoline skeleton specific to tetrodotoxin. The saxitoxin fraction consisted of neosaxitoxin, saxitoxin and two unknown compounds.
