标题：Expression cloning of a human α1,4-N-acetylglucosaminyltransferase that forms GlcNAcα1→4Galβ→R, a glycan specifically expressed in the gastric gland mucous cell-type mucin
期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：1999
卷号：96
期号：16
页码：8991-8996
DOI：10.1073/pnas.96.16.8991
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：Among mucus-secreting cells, the gastric gland mucous cells, Brunner's glands, accessory glands of pancreaticobiliary tract, and pancreatic ducts exhibiting gastric metaplasia are unique in that they express class III mucin identified by paradoxical Con A staining composed of periodate oxidation, sodium borohydride reduction, Con A, and horseradish peroxidase reaction. Recently it was shown that these mucous cells secrete glycoproteins having GlcNAc1[->]4Gal{beta}[->]R at nonreducing terminals of the carbohydrate moieties. Herein we describe the expression cloning of a cDNA encoding a human 1,4-N-acetylglucosaminyltransferase (4GnT), a key enzyme for the formation of GlcNAc1[->]4Gal{beta}1[->]R. COS-1 cells were thus cotransfected with a stomach cDNA library and a leukosialin cDNA. Transfected COS-1 cells were screened by using monoclonal antibodies specific for GlcNAc1[->]4Gal{beta}[->]R and enriched by fluorescence-activated cell sorting. Sibling selection of recovered plasmids resulted in a cDNA clone that directs the expression of GlcNAc1[->]4Gal{beta}[->]R. The deduced amino acid sequence predicts a type II membrane protein with 340 amino acids, showing no significant similarity with any other proteins. The 4GnT gene is located at chromosome 3p14.3, and its transcripts are expressed in the stomach and pancreas. An in vitro GlcNAc transferase assay by using a soluble 4GnT revealed that 1,4-linked GlcNAc residues are transferred most efficiently to core 2 branched O-glycans (Gal{beta}1[->]4GlcNAc{beta}1[->]6(Gal{beta}1[->]3)GalNAc), forming GlcNAc1[->]4Gal{beta}1[->]4GlcNAc{beta}1[->]6(GlcNAc1[->]4Gal{beta}1[->]3)GalNAc. Transfection of 4GnT cDNA into gastric adenocarcinoma AGS cells produced class III mucin, indicating that 4GnT is responsible for the formation of class III Con A reactivity. These results indicate that the 4GnT is a glycosyltransferase that forms 1,4-linked GlcNAc residues, preferentially in O-glycans.
