RESUMO
In this research, we firstly demonstrated that physcion, an anthraquinone derivative, specifically increased the expression of the human α2,8-sialyltransferase (hST8Sia VI) gene in SK-N-BE(2)-C human neuroblastoma cells. To establish the mechanism responsible for the up-regulation of hST8Sia VI gene expression in physcion-treated SK-N-BE(2)-C cells, the putative promoter region of the hST8Sia VI gene was functionally characterized. Promoter analysis with serially truncated fragments of the 5'-flanking region showed that the region between -320 and -240 is crucial for physcion-induced transcription of hST8Sia VI in SK-N-BE(2)-C cells. Putative binding sites for transcription factors Pax-5 and NF-Y are located at this region. The Pax-5 binding site at -262 to -256 was essential for the expression of the hST8Sia VI gene by physcion in SK-N-BE(2)-C cells. Moreover, the transcription of hST8Sia VI induced by physcion in SK-N-BE(2)-C cells was inhibited by extracellular signal-regulated protein kinase (ERK) inhibitor U0126 and p38 mitogen-activated protein kinase (MAPK) inhibitor SB203580, but not c-Jun N-terminal kinase (JNK) inhibitor SP600125. These results suggest that physcion upregulates hST8Sia VI gene expression via ERK and p38 MAPK pathways in SK-N-BE(2)-C cells.
Assuntos
Neoplasias Encefálicas/genética , Emodina/análogos & derivados , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neuroblastoma/genética , Sialiltransferases/genética , Regulação para Cima/efeitos dos fármacos , Região 5'-Flanqueadora/genética , Apoptose/efeitos dos fármacos , Apoptose/genética , Sequência de Bases , Neoplasias Encefálicas/enzimologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Sobrevivência Celular/genética , Emodina/química , Emodina/isolamento & purificação , Emodina/farmacologia , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Humanos , Neuroblastoma/enzimologia , Regiões Promotoras Genéticas , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sequência de DNA , Deleção de Sequência , Sialiltransferases/metabolismo , Ativação Transcricional/efeitos dos fármacos , Ativação Transcricional/genética , Regulação para Cima/genética , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismoRESUMO
In the present study, we firstly found that cordycepin elevated the gene expression of the human GD3 synthase (hST8Sia I) in human neuroblastoma SK-N-BE(2)-C cells. To elucidate the mechanism underlying the upregulation of hST8Sia I gene expression in cordycepin-treated SK-N-BE(2)-C cells, functional characterization of the promoter region of the hST8Sia I gene was performed. Analysis of promoter activity using varying lengths of 5'-flanking region showed a dramatic increase by cordycepin in the -1146 to -646 region, which contains putative binding sites for transcription factors c-Ets-1, CREB, AP-1, and NF-κB. Site-directed mutagenesis for these binding sites and chromatin immunoprecipitation assay revealed that the NF-κB binding site at -731 to -722 is essential for the cordycepin-induced expression of the hST8Sia I in SK-N-BE(2)-C cells. Moreover, the hST8Sia I expression induced by cordycepin was significantly repressed by pyrrolidinedithiocarbamate, an inhibitor of NF-κB. These results suggested that cordycepin induces upregulation of hST8Sia I gene expression through NF-κB activation in SK-N-BE(2)-C cells.