Transcription of the beta-galactoside alpha 2,6-sialyltransferase gene in B lymphocytes is directed by a separate and distinct promoter.
A single human gene, SIAT1, encodes the beta-galactoside alpha 2,6-sialyltransferase from which multiple mRNA isoforms are generated. In rat, expression of the hepatic mRNA isoform (Form 1) has been defined with respect to the transcriptional initiation site and promoter region. We show here that a similar hepatic SIAT1 mRNA isoform exists in human. Another human mRNA isoform, a mature B-cell-specific mRNA isoform (Form 2), was previously reported. Here, we used 5'-RACE and S1 nuclease protection analysis to define the 5'-untranslated region of Form 2 human SIAT1 mRNA. We demonstrate conclusively that Form 2 mRNA is initiated from a point completely distinct from that of Form 1 mRNA. A number of cis-acting regulatory elements residing immediately 5'of the Form 2 initiation site includes AP-1, AP-2, NF-kappa B, NF-IL6, C/EBP, and CREB. A TATAA box is also present 29 bp 5' of the transcriptional initiation site. CAT reporter gene expression from serially-truncated segments of the 5'-flanking region of the Form 2 initiation site indicates that the segment between -784 and +125 was sufficient to promote high level CAT expression in Louckes, a mature B-cell line. The 5'-flanking region to the human Form 1 initiation site is competent in expression of CAT upon transfection of the fusion construct into HepG2, a human hepatoma cell line. Cellular specificity of expression is apparently retained. Louckes cells expressed CAT efficiently from Form 2 promoter but only marginally from the Form 1 promoter. In contrast, CAT expression from Form 1 promoter is more efficient than from the Form 2 promoter in HepG2 cells.
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Department of Molecular and Cellular Biology, Roswell Park Cancer Institute, Buffalo, NY 14263, USA.
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JT
D000818:Animals; D001402:B-Lymphocytes / Q000166:cytology / Q000378:metabolism*; D001483:Base Sequence; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D004247:DNA / Q000235:genetics; D017930:Genes, Reporter; D006801:Humans; D008099:Liver / Q000378:metabolism; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D051381:Rats; D012799:Sialyltransferases / Q000235:genetics*; D014158:Transcription, Genetic; D014162:Transfection; D014407:Tumor Cells, Cultured; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase
D012333:RNA, Messenger; D004247:DNA; D015500:Chloramphenicol O-Acetyltransferase; D012799:Sialyltransferases; D000094802:beta-D-Galactoside alpha 2-6-Sialyltransferase