Isolation and functional analysis of the human glioblastoma-specific promoter region of the human GD3 synthase (hST8Sia I) gene

We identified the promoter region of the human GD3 synthase (hST8Sia I) gene to elucidate the mechanism underlying the regulation of hST8Sia I expression in human glioblastoma cells. The 5′-rapid amplification of cDNA end using mRNA prepared from U-87MG cells revealed the presence of transcription start site of hST8Sia I gene, and the 5′-terminal analysis of its product showed that transcription started from 648 nucleotides upstream of the translational initiation site. Functional analysis of the 5′-flanking region of the hST8Sia I gene by transient expression method revealed that the region from -638 to -498 is important for transcriptional activity of the hST8Sia I gene in U-87MG and T98G cells. This region lacks apparent TATA and CAAT boxes, but contains putative binding sites for transcription factors AREB6 and Elk-1. Site-directed mutagenesis and transient transfection assays demonstrated that both AREB6 and Elk-1 elements in this region were required for the promoter activity in U-87MG and T98G cells. These results indicated that both AREB6 and Elk-1 might play an essential role in the transcriptional activity of hST8Sia I gene essential for GD3 synthesis in human glioblastoma cells.