RESUMEN
Apios tuber lectin, named ATL, was isolated from Apios americana Medikus by two chromatography steps, hydrophobic chromatography and anion-exchange chromatography. The minimum concentration required for the hemagglutination activity toward rabbit erythrocytes of ATL was 4 µg/mL. ATL was composed of a homodimer of 28.4 kDa subunits. The amino acid sequence of ATL was similar to those of other legume lectins. The lectin showed moderate stability toward heating and acidic pH, and the binding affinity against several monosaccharides, such as D-glucosamine and D-galactosamine. ATL also bound to desialylated or agalactosylated glycoproteins such as asialo and agalacto transferrin. ATL decreased the transepithelial electrical resistance across human intestinal Caco-2 cell monolayers, suggesting the effect on the tight junction-mediated paracellular transport.
Asunto(s)
Fabaceae/química , Lectinas de Plantas/aislamiento & purificación , Tubérculos de la Planta/química , Secuencia de Aminoácidos , Animales , Células CACO-2 , Carbohidratos/análisis , Impedancia Eléctrica , Electroforesis en Gel de Poliacrilamida , Hemaglutinación/efectos de los fármacos , Humanos , Concentración de Iones de Hidrógeno , Iones , Metales/farmacología , Datos de Secuencia Molecular , Peso Molecular , Péptidos/química , Lectinas de Plantas/química , Lectinas de Plantas/farmacología , Conejos , Homología de Secuencia de Aminoácido , Glycine max/química , TemperaturaRESUMEN
Members of the enhancer of split- and hairy-related protein (SHARP) family, SHARP-1 and SHARP-2, are basic helix-loop-helix transcriptional repressors and belong to the clock genes. In this study, an effect of retinoic acid (RA) on the SHARP family gene expression in the differentiated cells was examined. RA rapidly and temporarily induced the SHARP-2 mRNA expression in hepatic H4IIE cells. Then, whether the SHARP-2 mRNA expression is altered by dexamethasone (Dex), insulin, and the combination of RA and Dex or RA and insulin was examined. Dex had different effects on the expression of SHARP-2 mRNA in the presence or absence of RA. Then, the molecular mechanisms were investigated using inhibitors of various signaling molecules. The RA-induction of SHARP-2 mRNA level was mainly inhibited by LY294002, staurosporine, and actinomycin D, respectively. Finally, whether RA acts on the transcriptional regulatory region of the SHARP-2 gene was analysed using luciferase reporter gene assay. At least two RA-responsive regions were mapped at the nucleotide sequences between -3,700 and -1,600 of the SHARP-2 gene. In addition, this effect was dependent on the RA receptor and retinoid X receptor. Thus, we conclude that RA stimulated transcription of the SHARP-2 gene via multiple pathways.