RESUMO
The function of the cystic fibrosis transmembrane conductance regulator (CFTR) as a Cl- channel in the apical membrane of epithelial cells is extensively documented. However, less is known about the molecular determinants of CFTR residence in the apical membrane, basal regulation of its Cl- channel activity, and its reported effects on the function of other transporters. These aspects of CFTR function likely require specific interactions between CFTR and unknown proteins in the apical compartment of epithelial cells. Here we report that CFTR interacts with the recently discovered protein, EBP50 (ERM-binding phosphoprotein 50). EBP50 is concentrated at the apical membrane in human airway epithelial cells, in vivo, and CFTR and EBP50 associate in in vitro binding assays. The CFTR-EBP50 interaction requires the COOH-terminal DTRL sequence of CFTR and utilizes either PDZ1 or PDZ2 of EBP50, although binding to PDZ1 is of greater affinity. Through formation of a complex, the interaction between CFTR and EBP50 may influence the stability and/or regulation of CFTR Cl- channel function in the cell membrane and provides a potential mechanism through which CFTR can affect the activity of other apical membrane proteins.
Assuntos
Proteínas de Transporte/metabolismo , Regulador de Condutância Transmembrana em Fibrose Cística/metabolismo , Fosfoproteínas/metabolismo , Trocadores de Sódio-Hidrogênio , Sequência de Aminoácidos , Técnicas Biossensoriais , Brônquios/citologia , Linhagem Celular , Proteínas do Citoesqueleto/metabolismo , Imunofluorescência , Humanos , Proteínas de Membrana/fisiologia , Dados de Sequência Molecular , Fragmentos de Peptídeos/metabolismo , Ligação Proteica/fisiologiaRESUMO
Cytoplasmic, nitric oxide-activated guanylate cyclases are expressed in many regions of the mammalian brain and are thought to participate in functions as diverse as synaptogenesis and long-term potentiation. In this study, we have characterized cytoplasmic guanylate cyclases in the nervous system of an invertebrate, the American lobster. Cytoplasmic cyclase specific activity is higher in lobster nerve cord than in any other lobster tissue tested, and considerably higher than in typical rat tissues (cerebellum, lung, and liver). However, nitric oxide donors have minimal effects on lobster nerve cord cyclic GMP production, when applied either to intact tissue or to cytoplasmic extracts. Parallel immunocytochemical studies, using an anti-cyclic GMP antibody, reveal that only a small subset of lobster neurons responds to nitric oxide with a significant elevation of cyclic GMP levels. HPLC analysis of nerve cord cytoplasm reveals two chromatographically separable cyclases, a minor nitric oxide-sensitive form whose retention time is identical to that of the conventional mammalian enzyme and a more abundant nitric oxide-insensitive form that appears to be novel. The physiological function and phylogenetic distribution of this nitric oxide-insensitive enzyme, and the signaling mechanisms that regulate its activity, are not known.
Assuntos
Citoplasma/enzimologia , Guanilato Ciclase/efeitos dos fármacos , Guanilato Ciclase/metabolismo , Óxido Nítrico/farmacologia , Animais , Cromatografia Líquida de Alta Pressão , Resistência a Medicamentos , Nephropidae , Sistema Nervoso/citologia , Sistema Nervoso/enzimologia , Neurônios/enzimologia , Nitroprussiato/farmacologiaRESUMO
BACKGROUND & AIMS: Guanylin, an endogenous gastrointestinal peptide, causes the translocation of NaCl from interstitial fluid to the intestinal lumen. The aim of this study was to examine whether changes in dietary salt intake lead to compensatory changes in expression of the guanylin signaling pathway. METHODS: Rats received low-, normal-, or high-sodium diets for 1 week. Colonic guanylin expression was evaluated by Western and Northern blotting, rates of guanylin secretion by measuring biologically active guanylin released into the medium from colon explants, and expression of the guanylin receptor (C-type guanylate cyclase) by Northern blotting and bioassay. RESULTS: By every criterion, the low-salt diet reduced expression of guanylin to 30%-40% of the level found in control animals. Guanylin receptor expression was also decreased, although less dramatically and with a lower statistical significance. For both guanylin and guanylin receptor, the high-salt diet had no significant effect on expression. CONCLUSIONS: The data support the hypothesis that the guanylin pathway is down-regulated as an adaptive response to salt restriction.
Assuntos
Colo/metabolismo , Hormônios Gastrointestinais , Peptídeos/metabolismo , Transdução de Sinais/efeitos dos fármacos , Cloreto de Sódio na Dieta/administração & dosagem , Animais , Regulação para Baixo , Masculino , Peptídeos Natriuréticos , Ratos , Ratos Sprague-DawleyAssuntos
Radioterapia/métodos , Tomografia Computadorizada por Raios X/métodos , Cateterismo/instrumentação , Meios de Contraste , Custos e Análise de Custo , Humanos , Fotografação , Radioterapia/economia , Radioterapia/instrumentação , Dosagem Radioterapêutica , Tecnologia Radiológica/economia , Tecnologia Radiológica/instrumentação , Fatores de Tempo , Tomografia Computadorizada por Raios X/economia , Tomografia Computadorizada por Raios X/instrumentaçãoRESUMO
Radioisotope myelography can be used effectively for the detection of spinal fluid leaks following surgical procedures involving the spine and spinal cord. The advantages of this approach are discussed in relation to currently used techniques and are illustrated by a case report.