RESUMO
Research in perception and attention has typically sought to evaluate cognitive mechanisms according to the average response to a manipulation. Recently, there has been a shift toward appreciating the value of individual differences and the insight gained by exploring the impacts of between-participant variation on human cognition. However, a recent study suggests that many robust, well-established cognitive control tasks suffer from surprisingly low levels of test-retest reliability (Hedge, Powell, & Sumner, 2018b). We tested a large sample of undergraduate students (n = 160) in two sessions (separated by 1-3 weeks) on four commonly used tasks in vision science. We implemented measures that spanned a range of perceptual and attentional processes, including motion coherence (MoCo), useful field of view (UFOV), multiple-object tracking (MOT), and visual working memory (VWM). Intraclass correlations ranged from good to poor, suggesting that some task measures are more suitable for assessing individual differences than others. VWM capacity (intraclass correlation coefficient [ICC] = 0.77), MoCo threshold (ICC = 0.60), UFOV middle accuracy (ICC = 0.60), and UFOV outer accuracy (ICC = 0.74) showed good-to-excellent reliability. Other measures, namely the maximum number of items tracked in MOT (ICC = 0.41) and UFOV number accuracy (ICC = 0.48), showed moderate reliability; the MOT threshold (ICC = 0.36) and UFOV inner accuracy (ICC = 0.30) showed poor reliability. In this paper, we present these results alongside a summary of reliabilities estimated previously for other vision science tasks. We then offer useful recommendations for evaluating test-retest reliability when considering a task for use in evaluating individual differences.
Assuntos
Atenção , Visão Ocular , Cognição/fisiologia , Humanos , Memória de Curto Prazo , Reprodutibilidade dos TestesRESUMO
Over the last two decades, important contributions were made at national, European and international levels to foster collaboration into rare diseases research. The European Union (EU) has put much effort into funding rare diseases research, encouraging national funding organizations to collaborate together in the E-Rare program, setting up European Reference Networks for rare diseases and complex conditions, and initiating the International Rare Diseases Research Consortium (IRDiRC) together with the National Institutes of Health in the USA. Co-ordination of the activities of funding agencies, academic researchers, companies, regulatory bodies, and patient advocacy organizations and partnerships with, for example, the European Research Infrastructures maximizes the collective impact of global investments in rare diseases research. This contributes to accelerating progress, for example, in faster diagnosis through enhanced discovery of causative genes, better understanding of natural history of rare diseases through creation of common registries and databases and boosting of innovative therapeutic approaches. Several examples of funded pre-clinical and clinical gene therapy projects show that integration of multinational and multidisciplinary expertize generates new knowledge and can result in multicentre gene therapy trials. International collaboration in rare diseases research is key to improve the life of people living with a rare disease.
Assuntos
Pesquisa Biomédica/organização & administração , Cooperação Internacional , Doenças Raras/terapia , Pesquisa Biomédica/economia , União Europeia , Humanos , Doenças Raras/diagnósticoRESUMO
The coupling between the 'coffee-ring' effect and liquid-liquid phase separation is examined for ternary mixtures of solvent, polymer and semiconductor nanocrystal. Specifically, we study mixtures of toluene, polystyrene (PS) and colloidal silicon nanocrystals (SiNCs) using real-space imaging and spectroscopic techniques to resolve the kinetic morphology of the drying front for varied molecular weight of the PS. Our results demonstrate that the size of the polymer has a significant impact on both phase-separation and drying, and we relate these observations to simulations, measured and predicted binodal curves, and the observed shape of the flow field at the contact line. The results inform a deposition process that reduces the influence of drying instabilities for low-molecular-weight polymers while paving the way for more detailed and generic computational descriptions of drying instabilities in complex fluids.
Assuntos
Nanopartículas/química , Polímeros/química , Cinética , Transição de Fase , Poliestirenos/química , Propriedades de SuperfícieRESUMO
DISC1 (Disrupted-In-Schizophrenia 1) has been associated with schizophrenia in multiple genetic studies. Studies from our laboratory have shown that Disc1, the mouse ortholog of DISC1, is highly expressed in the dentate gyrus of the hippocampus in the adult mouse brain. Because developmental dysfunction of the hippocampus is thought to play a major role in schizophrenia pathogenesis, and the dentate gyrus is a major locus for adult neurogenesis in the mouse, we investigated Disc1 expression during mouse brain development. Strikingly, Disc1 is strongly expressed in the hippocampus during all stages of hippocampal development, from embryonic day 14 through adulthood. Disc1 mRNA was detected in the dentate gyrus at all stages in which this structure was identifiable, as well as in the cornu ammonis (CA) fields of the hippocampus, the subiculum and adjacent entorhinal cortex, and the developing cerebral neocortex, hypothalamus, and olfactory bulbs, all of which also express Disc1 in the adult mouse brain. In addition, Disc1 mRNA was seen in regions of the developing mouse brain which do not express Disc1 during adulthood, regions including the bed nucleus of the stria terminalis, reticular thalamic nucleus and reuniens thalamic nucleus. These results demonstrate that Disc1 marks the hippocampus from its earliest stages, and suggest that developmental Disc1 dysfunction may lead to defects in hippocampal function that are associated with schizophrenia.
Assuntos
Regulação da Expressão Gênica no Desenvolvimento , Hipocampo/embriologia , Hipocampo/fisiologia , Proteínas do Tecido Nervoso/genética , Animais , Córtex Cerebral/embriologia , Córtex Cerebral/fisiologia , Feminino , Hibridização In Situ , Camundongos , Gravidez , RNA Mensageiro/análise , Tálamo/embriologia , Tálamo/fisiologiaRESUMO
BACKGROUND: Cysteinyl leukotrienes (CysLTs) are bioactive lipids that have been shown to contribute to allergic and inflammatory diseases. Eosinophils and mast cells have the capacity to produce large amounts of CysLTs after allergic or non-allergic stimulation. Molecular identification of both the synthetic and signalling proteins in the CysLT pathway allows the investigation of expression of the CysLT enzymes and receptors in active allergic rhinitis. OBJECTIVE: We examined the expression of the proteins involved in the synthesis of CysLTs and the cysteinyl leukotriene-1 (CysLT1) and cysteinyl leukotriene-2 (CysLT2) receptors in inflammatory cells from patients with active seasonal allergic rhinitis. METHODS: Nasal lavage samples were obtained from patients during active seasonal allergic rhinitis. Specific cellular immunocytochemical techniques were used to detect the cysteinyl leukotriene synthetic proteins, namely 5-lipoxygenase (5-LO), 5-lipoxygenase-activating protein (FLAP) and leukotriene C4 synthase (LTC4S). In situ hybridization and immunocytochemical techniques were used to identify the mRNA and proteins for the CysLT1 and CysLT2 receptors. RESULTS: 5-LO, FLAP and LTC4S, and the CysLT1 and CysLT2 receptors were expressed in the majority of eosinophils and in subsets of mast cells and mononuclear cells. 5-LO, FLAP and the CysLT1 receptor, but not LTC4S or the CysLT2 receptor, were expressed in a subset of nasal neutrophils. CONCLUSIONS: Our study demonstrates the presence of CysLT pathway proteins in key allergic and inflammatory cells from the upper airway of patients with active seasonal allergic rhinitis. Our expression data highlight the potential of CysLT-modifying agents to treat both upper and lower airway symptoms in patients suffering from allergic rhinitis and asthma.
Assuntos
Cisteína/biossíntese , Leucotrienos/biossíntese , Rinite Alérgica Sazonal/metabolismo , Proteínas Ativadoras de 5-Lipoxigenase , Adulto , Araquidonato 5-Lipoxigenase/metabolismo , Proteínas de Transporte/metabolismo , Cisteína/genética , Expressão Gênica , Glutationa Transferase/metabolismo , Humanos , Hibridização In Situ , Leucotrienos/genética , Proteínas de Membrana/metabolismo , Pessoa de Meia-Idade , Líquido da Lavagem Nasal/química , RNA Mensageiro/genética , Receptores de Leucotrienos/biossíntese , Receptores de Leucotrienos/genética , Transdução de SinaisRESUMO
Bipolar affective disorder is one of the most common mental illnesses with a population prevalence of approximately 1%. The disorder is genetically complex, with an increasing number of loci being implicated through genetic linkage studies. However, the specific genetic variations and molecules involved in bipolar susceptibility and pathogenesis are yet to be identified. Genetic linkage analysis has identified a bipolar disorder susceptibility locus on chromosome 4q35, and the interval harbouring this susceptibility gene has been narrowed to a size that is amenable to positional cloning. We have used the resources of the Human Genome Project (HGP) and Celera Genomics to identify overlapping sequenced BAC clones and sequence contigs that represent the region implicated by linkage analysis. A combination of bioinformatic tools and laboratory techniques have been applied to annotate this DNA sequence data and establish a comprehensive transcript map that spans approximately 5.5 Mb. This map encompasses the chromosome 4q35 bipolar susceptibility locus, which localises to a "most probable" candidate interval of approximately 2.3 Mb, within a more conservative candidate interval of approximately 5 Mb. Localised within this map are 11 characterised genes and eight novel genes of unknown function, which together provide a collection of candidate transcripts that may be investigated for association with bipolar disorder. Overall, this region was shown to be very gene-poor, with a high incidence of pseudogenes, and redundant and novel repetitive elements. Our analysis of the interval has demonstrated a significant difference in the extent to which the current HGP and Celera sequence data sets represent this region.
Assuntos
Transtorno Bipolar/genética , Mapeamento Cromossômico , Cromossomos Humanos Par 4 , Ligação Genética , Predisposição Genética para Doença/genética , HumanosRESUMO
Two classes of cysteinyl leukotriene receptor, CysLT(1) and CysLT(2), have been identified and pharmacologically characterized in human tissues. Although the CysLT(1) receptor mediates the proinflammatory effects of leukotrienes in human asthma, the physiological roles of CysLT(2) receptor are not defined, and a suitable mouse model would be useful in delineating function. We report here the molecular cloning and characterization of the mouse CysLT(2) receptor (mCysLT(2)R) from heart tissue. mCysLT(2)R cDNA encodes a protein of 309 amino acids, truncated at both ends compared with the human ortholog (hCysLT(2)R). The gene resides on the central region of mouse chromosome 14 and is composed of 6 exons with the entire coding region located in the last exon. Two 5'-untranslated region splice variants were identified with the short form lacking exon 3 as the predominant transcript. Although the overall expression of mCysLT(2)R is very low, the highest expression was detected in spleen, thymus, and adrenal gland by ribonuclease protection assay, and discrete sites of expression in heart were observed by in situ hybridization. Intracellular calcium mobilization in response to cysteinyl leukotriene administration was detected in human embryonic kidney 293T cells transfected with recombinant mCysLT(2)R with a rank order of potency leukotriene C(4)(LTC(4) ) = LTD(4)>>LTE(4). [(3)H]LTD(4) binding to membranes expressing mCysLT(2)R could be effectively competed by LTC(4) and LTD(4) and only partially inhibited by LTE(4) and BAYu9773. The identification of mCysLT(2)R will be useful for establishing CysLT(2)R-deficient mice and determining novel leukotriene functions.
Assuntos
Processamento Alternativo , DNA Complementar/metabolismo , Proteínas de Membrana , Receptores de Leucotrienos/química , Receptores de Leucotrienos/genética , Regiões 5' não Traduzidas , Glândulas Suprarrenais/metabolismo , Sequência de Aminoácidos , Animais , Sequência de Bases , Ligação Competitiva , Northern Blotting , Cálcio/metabolismo , Linhagem Celular , Células Cultivadas , Mapeamento Cromossômico , Clonagem Molecular , Cruzamentos Genéticos , Relação Dose-Resposta a Droga , Éxons , Humanos , Hibridização In Situ , Íntrons , Leucotrieno C4/metabolismo , Leucotrieno D4/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Modelos Genéticos , Dados de Sequência Molecular , Miocárdio/metabolismo , Estrutura Terciária de Proteína , RNA Mensageiro/metabolismo , Ensaio Radioligante , Proteínas Recombinantes/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Homologia de Sequência de Aminoácidos , Baço/metabolismo , Timo/metabolismo , Distribuição Tecidual , TransfecçãoRESUMO
The two mammalian neuropeptides NPFF and NPAF have been shown to have important roles in nociception, anxiety, learning and memory, and cardiovascular reflex. Two receptors (FF1 and FF2) have been molecularly identified for NPFF and NPAF. We have now characterized a novel gene designated NPVF that encodes two neuropeptides highly similar to NPFF. NPVF mRNA was detected specifically in a region between the dorsomedial and ventromedial hypothalamic nuclei. NPVF-derived peptides displayed higher affinity for FF1 than NPFF-derived peptides, but showed poor agonist activity for FF2. Following intracerebral ventricular administration, a NPVF-derived peptide blocked morphine-induced analgesia more potently than NPFF in both acute and inflammatory models of pain. In situ hybridization analysis revealed distinct expression patterns of FF1 and FF2 in the rat central nervous system. FF1 was broadly distributed, with the highest levels found in specific regions of the limbic system and the brainstem where NPVF-producing neurons were shown to project. FF2, in contrast, was mostly expressed in the spinal cord and some regions of the thalamus. These results indicate that the endogenous ligands for FF1 and FF2 are NPVF- and NPFF-derived peptides, respectively, and suggest that the NPVF/FF1 system may be an important part of endogenous anti-opioid mechanism.
Assuntos
Analgésicos/farmacologia , Morfina/farmacologia , Oligopeptídeos/isolamento & purificação , Sequência de Aminoácidos , Analgésicos/antagonistas & inibidores , Analgésicos Opioides/antagonistas & inibidores , Analgésicos Opioides/farmacologia , Animais , Células CHO , Células Cultivadas , Sistema Nervoso Central , Cricetinae , Modelos Animais de Doenças , Interações Medicamentosas , Humanos , Masculino , Camundongos , Dados de Sequência Molecular , Morfina/antagonistas & inibidores , Oligopeptídeos/genética , Oligopeptídeos/farmacologia , Dor/tratamento farmacológico , Medição da Dor/efeitos dos fármacos , Ratos , Ratos Sprague-Dawley , Homologia de Sequência de AminoácidosRESUMO
We have isolated and determined the coding sequences of human and mouse orthologs of the rat orphan G-protein-coupled receptor GPR54. Mouse and rat GPR54 are nearly 95% identical to each other, and both are approximately 85% identical to human GPR54 at the amino acid level. Screening of agonists for GPR54 identified several invertebrate neuropeptides of the RFamide and RWamide family that were able to activate GPR54 at microM range through the G(alpha)q pathway. Substitution analysis showed that the C-terminal optimal sequence of GPR54-activating peptides is Gly-Leu-Arg-Trp-NH2. Northern analysis of human GPR54 detected expression in several peripheral tissues and many regions of the central nervous system.
Assuntos
Neuropeptídeos/metabolismo , Receptores de Neuropeptídeos/genética , Sequência de Aminoácidos , Animais , Clonagem Molecular , DNA Complementar/análise , Relação Dose-Resposta a Droga , FMRFamida/química , Expressão Gênica , Humanos , Camundongos , Dados de Sequência Molecular , Neuropeptídeos/farmacologia , Ratos , Receptores Acoplados a Proteínas G , Receptores de Kisspeptina-1 , Receptores de Neuropeptídeos/agonistas , Receptores de Neuropeptídeos/isolamento & purificação , Receptores de Neuropeptídeos/metabolismo , Homologia de Sequência de AminoácidosRESUMO
Melanin-concentrating hormone (MCH) is a 19-aa cyclic neuropeptide originally isolated from chum salmon pituitaries. Besides its effects on the aggregation of melanophores in fish several lines of evidence suggest that in mammals MCH functions as a regulator of energy homeostasis. Recently, several groups reported the identification of an orphan G protein-coupled receptor as a receptor for MCH (MCH-1R). We hereby report the identification of a second human MCH receptor termed MCH-2R, which shares about 38% amino acid identity with MCH-1R. MCH-2R displayed high-affinity MCH binding, resulting in inositol phosphate turnover and release of intracellular calcium in mammalian cells. In contrast to MCH-1R, MCH-2R signaling is not sensitive to pertussis toxin and MCH-2R cannot reduce forskolin-stimulated cAMP production, suggesting an exclusive G(alpha)q coupling of the MCH-2R in cell-based systems. Northern blot and in situ hybridization analysis of human and monkey tissue shows that expression of MCH-2R mRNA is restricted to several regions of the brain, including the arcuate nucleus and the ventral medial hypothalamus, areas implicated in regulation of body weight. In addition, the human MCH-2R gene was mapped to the long arm of chromosome 6 at band 6q16.2-16.3, a region reported to be associated with cytogenetic abnormalities of obese patients. The characterization of a second mammalian G protein-coupled receptor for MCH potentially indicates that the control of energy homeostasis in mammals by the MCH neuropeptide system may be more complex than initially anticipated.
Assuntos
Encéfalo/metabolismo , Cromossomos Humanos Par 22 , Receptores do Hormônio Hipofisário/genética , Receptores do Hormônio Hipofisário/metabolismo , Receptores do Hormônio Hipofisário/fisiologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Células CHO , Células COS , Membrana Celular/fisiologia , Chlorocebus aethiops , Mapeamento Cromossômico , Cricetinae , Feminino , Humanos , Hibridização In Situ , Masculino , Dados de Sequência Molecular , Oncorhynchus keta , Especificidade de Órgãos , Hipófise/química , Hipófise/fisiologia , Ensaio Radioligante , Receptores Acoplados a Proteínas G , Receptores do Hormônio Hipofisário/química , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , TransfecçãoRESUMO
Differences between benign prostatic hyperplasia (BPH) and normal prostate tissue at the level of mRNA expression provide an opportunity to identify candidate genes for this disease. A cDNA subtraction procedure was used to isolate differentially expressed genes in BPH. The subtraction was done by solution hybridization of BPH cDNA against excess normal prostate cDNA. We identified known, EST, and novel genes by sequence and database analysis of the subtracted cDNAs. Several of these cDNAs were used as probes in Northern blotting analysis to confirm over-expression of their corresponding mRNAs in BPH tissues. One highly upregulated sequence of interest shared identity with a known mRNA encoding human NELL2, a protein containing epidermal growth factor-like domains. NELL2 was not previously reported to be expressed in prostate and may code for a novel prostatic growth factor. In situ hybridization analysis of hyperplastic prostate specimens demonstrated that NELL2 mRNA expression is predominantly localized in basal cells of the epithelium. Disease-related changes in the levels of NELL2 may contribute to alterations in epithelial-stromal homeostasis in BPH. (J Histochem Cytochem 49:669-670, 2001)
Assuntos
Hiperplasia Prostática/metabolismo , RNA Mensageiro/metabolismo , Northern Blotting , DNA Complementar/análise , Regulação da Expressão Gênica , Humanos , Hibridização In Situ , Lactoferrina/genética , Lactoferrina/metabolismo , Masculino , Proteínas do Tecido Nervoso/genética , Proteínas do Tecido Nervoso/metabolismoRESUMO
The cysteinyl leukotrienes (CysLTs) are important mediators of human asthma. Pharmacologic and clinical studies show that the CysLTs exert most of their bronchoconstrictive and proinflammatory effects through activation of a putative, 7-transmembrane domain, G-protein-coupled receptor, the CysLT1 receptor. The initial molecular characterization of the CysLT1 receptor showed by in situ hybridization, the presence of CysLT1 receptor messenger RNA (mRNA) in human lung smooth-muscle cells and lung macrophages. We confirmed the results of these in situ hybridization analyses for the CysLT1 receptor, and produced the first immunohistochemical characterization of the CysLT1 receptor protein in human lung. The identification of the CysLT1 receptor in the lung is consistent with the antibronchoconstrictive and antiinflammatory actions of CysLT1 receptor antagonists. We also report the expression of CysLT1 receptor mRNA and protein in most peripheral blood eosinophils and pregranulocytic CD34+ cells, and in subsets of monocytes and B lymphocytes.
Assuntos
Leucócitos/metabolismo , Proteínas de Membrana , Receptores de Leucotrienos/biossíntese , Sangue , Humanos , Pulmão/imunologia , Receptores de Leucotrienos/análiseRESUMO
Stargardt-like macular dystrophy (STGD3, MIM 600110) and autosomal dominant macular dystrophy (adMD) are inherited forms of macular degeneration characterized by decreased visual acuity, macular atrophy and extensive fundus flecks. Genetic mapping data suggest that mutations in a single gene may be responsible for both conditions, already known to bear clinical resemblance. Here we limit the minimum genetic region for STGD3 and adMD to a 0.6-cM interval by recombination breakpoint mapping and identify a single 5-bp deletion within the protein-coding region of a new retinal photoreceptor-specific gene, ELOVL4, in all affected members of STGD3 and adMD families. Bioinformatic analysis of ELOVL4 revealed that it has homology to a group of yeast proteins that function in the biosynthesis of very long chain fatty acids. Our results are therefore the first to implicate the biosynthesis of fatty acids in the pathogenesis of inherited macular degeneration.
Assuntos
Proteínas do Olho/genética , Genes Dominantes/genética , Degeneração Macular/genética , Proteínas de Membrana/genética , Deleção de Sequência/genética , Sequência de Aminoácidos , Animais , Sequência de Bases , Mapeamento Cromossômico , Cromossomos Humanos Par 6/genética , Clonagem Molecular , Análise Mutacional de DNA , Éxons/genética , Proteínas do Olho/química , Proteínas do Olho/metabolismo , Feminino , Humanos , Hibridização In Situ , Íntrons/genética , Escore Lod , Macaca mulatta/genética , Degeneração Macular/patologia , Masculino , Proteínas de Membrana/química , Proteínas de Membrana/metabolismo , Camundongos , Dados de Sequência Molecular , Linhagem , RNA Mensageiro/análise , RNA Mensageiro/genética , Retina/metabolismo , Retina/patologia , Alinhamento de SequênciaRESUMO
A novel human Kir5.1 (inward rectifier K+ channel subunit, gene name KCNJ16) was identified through database searches. This human KCNJ16 was mapped to chromosome 17q25. The full-length cDNA was identified and its genomic structure was determined. Tissue distribution studies showed that human KCNJ16 is significantly expressed in human kidney, pancreas and thyroid gland. In situ hybridization revealed expression in convoluted tubule cells of kidney and in the acinar and ductal cells of pancreas. These suggest that human Kir5.1 may be involved in the regulation of fluid and pH balance, thus making it a potential therapeutic target for hypertension, renal failure, or pancreatic disease.
Assuntos
Cromossomos Humanos Par 17/genética , Rim/metabolismo , Pâncreas/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização , Canais de Potássio/genética , Sequência de Aminoácidos , Animais , Bases de Dados como Assunto , Etiquetas de Sequências Expressas , Humanos , Hibridização In Situ , Rim/citologia , Dados de Sequência Molecular , Pâncreas/citologia , Canais de Potássio/química , RNA Mensageiro/análise , RNA Mensageiro/genética , Mapeamento de Híbridos Radioativos , Alinhamento de SequênciaRESUMO
LRP5 is a novel member of the low-density lipoprotein receptor family that is genetically associated with Type 1 diabetes. As a start to defining the normal function of LRP5 and to generate testable hypotheses of its potential role in Type 1 diabetes pathogenesis, we carried out an extensive expression analysis of this gene at the mRNA and protein levels in normal human, monkey, and mouse, as well as in non-obese diabetic (NOD) mice at several stages of diabetes development. In all species, expression of LRP5 was found in four functionally important cell types: the distributed mononuclear phagocyte system, the islets of Langerhans, vitamin A-metabolizing cells, and CNS neurons. Given the critical role of macrophages in the onset and progression of islet cell destruction in Type 1 diabetes and the hypothesized role of retinoids as modifiers of diabetes progression, these findings suggest that LRP5 may confer Type 1 diabetes risk by altering the normal functioning of one or more of these regulatory systems. Specifically, given that the LRP5 polymorphisms associated with diabetes are in the promoter region of the gene, alterations in LRP5 expression may be responsible for diabetes susceptibility and therefore may be potential targets for therapeutic intervention. (J Histochem Cytochem 48:1357-1368, 2000)
Assuntos
Diabetes Mellitus Tipo 1/metabolismo , Ilhotas Pancreáticas/metabolismo , Macrófagos/metabolismo , Receptores de LDL/metabolismo , Vitamina A/metabolismo , Animais , Western Blotting , Linhagem Celular , Chlorocebus aethiops , Humanos , Imuno-Histoquímica , Hibridização In Situ , Túbulos Renais/metabolismo , Proteínas Relacionadas a Receptor de LDL , Fígado/citologia , Fígado/metabolismo , Proteína-5 Relacionada a Receptor de Lipoproteína de Baixa Densidade , Macaca mulatta , Camundongos , Camundongos Endogâmicos NOD , Neurônios/metabolismo , Epitélio Pigmentado Ocular/metabolismo , Baço/citologia , Baço/metabolismo , Timo/citologia , Timo/metabolismoRESUMO
Neuromedin U (NMU) is a neuropeptide with potent activity on smooth muscle which was isolated first from porcine spinal cord and later from other species. It is widely distributed in the gut and central nervous system. Peripheral activities of NMU include stimulation of smooth muscle, increase of blood pressure, alteration of ion transport in the gut, control of local blood flow and regulation of adrenocortical function. An NMU receptor has not been molecularly identified. Here we show that the previously described orphan G-protein-coupled receptor FM-3 (ref. 15) and a newly discovered one (FM-4) are cognate receptors for NMU. FM-3, designated NMU1R, is abundantly expressed in peripheral tissues whereas FM-4, designated NMU2R, is expressed in specific regions of the brain. NMU is expressed in the ventromedial hypothalamus in the rat brain, and its level is significantly reduced following fasting. Intracerebroventricular administration of NMU markedly suppresses food intake in rats. These findings provide a molecular basis for the biochemical activities of NMU and may indicate that NMU is involved in the central control of feeding.
Assuntos
Comportamento Alimentar/fisiologia , Proteínas de Membrana , Neuropeptídeos/metabolismo , Receptores de Superfície Celular/fisiologia , Receptores de Neurotransmissores/fisiologia , Sequência de Aminoácidos , Animais , Encéfalo/metabolismo , Linhagem Celular , Clonagem Molecular , Jejum , Humanos , Ligantes , Camundongos , Dados de Sequência Molecular , Obesidade/metabolismo , RNA Mensageiro/metabolismo , Ensaio Radioligante , Ratos , Receptores de Superfície Celular/análise , Receptores de Neurotransmissores/análise , Alinhamento de Sequência , Distribuição TecidualRESUMO
The contractile and inflammatory actions of the cysteinyl leukotrienes (CysLTs), LTC(4), LTD(4), and LTE(4), are thought to be mediated through at least two distinct but related CysLT G protein-coupled receptors. The human CysLT(1) receptor has been recently cloned and characterized. We describe here the cloning and characterization of the second cysteinyl leukotriene receptor, CysLT(2), a 346-amino acid protein with 38% amino acid identity to the CysLT(1) receptor. The recombinant human CysLT(2) receptor was expressed in Xenopus oocytes and HEK293T cells and shown to couple to elevation of intracellular calcium when activated by LTC(4), LTD(4), or LTE(4). Analyses of radiolabeled LTD(4) binding to the recombinant CysLT(2) receptor demonstrated high affinity binding and a rank order of potency for competition of LTC(4) = LTD(4) LTE(4). In contrast to the dual CysLT(1)/CysLT(2) antagonist, BAY u9773, the CysLT(1) receptor-selective antagonists MK-571, montelukast (Singulair(TM)), zafirlukast (Accolate(TM)), and pranlukast (Onon(TM)) exhibited low potency in competition for LTD(4) binding and as antagonists of CysLT(2) receptor signaling. CysLT(2) receptor mRNA was detected in lung macrophages and airway smooth muscle, cardiac Purkinje cells, adrenal medulla cells, peripheral blood leukocytes, and brain, and the receptor gene was mapped to chromosome 13q14, a region linked to atopic asthma.
Assuntos
Cisteína , Leucotrienos/metabolismo , Proteínas de Membrana , Receptores de Leucotrienos/genética , Receptores de Leucotrienos/metabolismo , Medula Suprarrenal/química , Clonagem Molecular , Humanos , Antagonistas de Leucotrienos/farmacologia , Leucotrieno C4/metabolismo , Leucotrieno D4/metabolismo , Leucotrieno E4/metabolismo , Pulmão/química , Modelos Moleculares , Miocárdio/química , Receptores de Leucotrienos/sangue , Proteínas Recombinantes/metabolismo , SRS-A/análogos & derivados , SRS-A/farmacologia , Análise de Sequência de DNA , Homologia de Sequência de Aminoácidos , Distribuição TecidualRESUMO
We report here a characterization of two families of calcium-activated K(+) channel beta-subunits, beta2 and beta3, which are encoded by distinct genes that map to 3q26.2-27. A single beta2 family member and four alternatively spliced variants of beta3 were investigated. These subunits have predicted molecular masses of 27. 1-31.6 kDa, share approximately 30-44% amino acid identity with beta1, and exhibit distinct but overlapping expression patterns. Coexpression of the beta2 or beta3a-c subunits with a BK alpha-subunit altered the functional properties of the current expressed by the alpha-subunit alone. The beta2 subunit rapidly and completely inactivated the current and shifted the voltage dependence for activation to more polarized membrane potentials. In contrast, coexpression of the beta3a-c subunits resulted in only partial inactivation of the current, and the beta3b subunit conferred an apparent inward rectification. Furthermore, unlike the beta1 and beta2 subunits, none of the beta3 subunits increased channel sensitivity to calcium or voltage. The tissue-specific expression of these beta-subunits may allow for the assembly of a large number of distinct BK channels in vivo, contributing to the functional diversity of native BK currents.
Assuntos
Cálcio/metabolismo , Canais de Potássio/genética , Processamento Alternativo , Sequência de Aminoácidos , Mapeamento Cromossômico , Cromossomos Humanos Par 3 , Clonagem Molecular , DNA Complementar , Humanos , Hibridização in Situ Fluorescente , Dados de Sequência Molecular , Canais de Potássio/química , Canais de Potássio/metabolismo , Homologia de Sequência de AminoácidosRESUMO
Fas-mediated apoptosis is an important regulator of cell survival, and abnormalities in this system have been shown to result in a number of human pathological conditions. A secreted member of the tumor necrosis factor receptor superfamily, DcR3, was recently reported to be amplified in human lung and colon cancers as a negative regulator of Fas-mediated apoptosis. We identified this gene, which we call M68. M68 genomic DNA, mRNA, and protein levels were examined in a series of human gastrointestinal tract tumors. Using M68 immunohistochemistry and a scoring system similar to that used for HER-2/neu, we found that M68 protein was overexpressed in 30 of 68 (44%) human adenocarcinomas of the esophagus, stomach, colon, and rectum. Tumors examined by Northern blot revealed M68 mRNA highly elevated in a similar fraction of primary tumors from the same gastrointestinal tract regions, as well as in the colon adenocarcinoma cell lines SW480 and SW1116. Further, we found M68 protein to be overexpressed in a substantial number of tumors in which gene amplification could not be detected by fluorescence in situ hybridization or quantitative genomic PCR, suggesting that overexpression of M68 may precede amplification in tumors. Finally, we find that M68 lies within a four-gene cluster that includes a novel helicase-like gene (NHL) related to RAD3/ERCC2, a plasma membrane Ras-related GTPase and a member of the stathmin family, amplification or overexpression of which may also contribute to cell growth and tumor progression.