Realtime Access Map
Molecular analysis of mammalian Neu4 sialidase gene promoter
There are four different mammalian sialidases that have been described; lysosomal (Neu1), cytoplasmic (Neu2), plasma membrane (Neu3), lysosomal/mitochondrial (Neu4). The activity of sialidase Neu4 enzyme against sialic acid containing ganglioside GM2 has been demonstrated. Biological role of sialidase Neu4 enzyme has been shown by the transfection of neuroglia cells from a Tay-Sachs patient with a Neu4-expressing plasmid showed clearance of accumulated ganglioside GM2. It has been also shown that sialidase Neu4 enzyme is responsible for degradation reactions of another ganglioside such as GD1a in brains of Neu4-/- mice. Aim of our study is to identify minimal promoter region of human Neu4 gene and demonstrate binding of transcription factors to this region. In our study, we used bioinformatic approaches to predict the sequence motifs where several specific transcription factors bind using TESS (Transcription Element Seach System) tool. We amplified seven different DNA fragments from human Neu4 promoter region, cloned into luciferase expression vector and performed reporter assay. We also performed electrophoretic mobility shift assay to demonstrate binding of transcription factors to candidate promoter region. We demonstrated that 187 bp upstream of Neu4 gene is minimal promoter region to control transcription from Neu4 gene. Electrophoretic mobility shift assay showed that 187 bp upstream region recruits several transcription factors. Our results demonstrated the minimal promoter region revealing several putative transcription factors such as Sp-1 and c-myc which might be responsible mainly for regulation of Neu4 gene transcription. The data we obtained might be useful to discover small molecules which can control Neu4 gene expression. High expression of Neu4 gene might be controlled using drugs or small molecules and the accumulated GM2 ganglioside in lysosomes of Tay-Sachs patients can be reduced.