RESUMO

Morquio A is an autosomal recessive disease caused by the deficiency of N-acetylgalactosamine-6-sulfate sulfatase (GALNS), leading to the lysosomal accumulation of keratan-sulfate and chondroitin-6-sulfate. We evaluated in HEK293 cells the effect of the cytomegalovirus immediate early enhancer/promoter (CMV) or the elongation factor 1alpha (EF1alpha) promoters, and the coexpression with the sulfatase modifying factor 1 (SUMF1) on GALNS activity. Four days postransfection GALNS activity in transfected cells with CMV-pIRES-GALNS reached a plateau, whereas in cells transfected with EF1alpha-pIRES-GALNS continued to increase until day 8. Co-transfection with pCXN-SUMF1 showed an increment up to 2.6-fold in GALNS activity. Finally, computational analysis of transcription factor binding-sites and CpG islands showed that EF1alpha promoter has long CpG islands and high-density binding-sites for Sp1 compared to CMV. These results show the advantage of the SUMF1 coexpression on GALNS activity and indicate a considerable effect on the expression stability using EF1alpha promoter compared to CMV.

Assuntos

Condroitina Sulfatases/metabolismo , Expressão Gênica , Fator 1 de Elongação de Peptídeos/genética , Regiões Promotoras Genéticas/genética , Sulfatases/metabolismo , Sítios de Ligação , Linhagem Celular , Condroitina Sulfatases/genética , Biologia Computacional , Ilhas de CpG/genética , Citomegalovirus/genética , Humanos , Oxirredutases atuantes sobre Doadores de Grupo Enxofre , Plasmídeos/genética , Fator de Transcrição Sp1/metabolismo , Transfecção