We describe transcriptional regulation of the human Gal beta1,4(3) GlcNAc alpha2,3-sialyltransferase VI (hST3Gal VI) gene. The 5'-RACE results indicated that two mRNA forms differ only in the 5'-untranslated region (types 1 and 2). The genomic structure shows that the transcriptional regulation of type 1 and type 2 mRNA depends on the P1 and P2 promoters, respectively. Northern blots of RNA derived from various human tissues showed that the expression level of type 2 mRNA is higher than that of type I in the prostate. To elucidate the molecular basis of hST3Gal VI gene ex. pression, we isolated and functionally characterized the genomic region containing the P1 and P2 promoters of hST3Gal VI. The activity of the P2 promoter is much higher than that of the P1 promoter in the prostate adenocarcinoma cell line PC-3. The results suggested that the hST3Gal VI gene is expressed specifically by alternative promoter utilization and is regulated in a tissue-restricted fashion at the level of transcription.