beta 1,3-Galactosyltransferase beta 3Gal-T5 acts on the GlcNAcbeta 1-->3Galbeta 1-->4GlcNAcbeta 1-->R sugar chains of carcinoembryonic antigen and other N-linked glycoproteins and is down-regulated in colon adenocarcinomas

J Biol Chem. 2001 Feb 2;276(5):3564-73. doi: 10.1074/jbc.M006662200. Epub 2000 Oct 31.

Abstract

We attempted to determine whether beta1,3-galactosyltransferase beta3Gal-T5 is involved in the biosynthesis of a specific subset of type 1 chain carbohydrates and expressed in a cancer-associated manner. We transfected Chinese hamster ovary (CHO) cells expressing Fuc-TIII with beta3Gal-T cDNAs and studied the relevant glycoconjugates formed. beta3Gal-T5 directs synthesis of Lewis type 1 antigens in CHO cells more efficiently than beta3Gal-T1, whereas beta3Gal-T2, -T3, and -T4 are almost unable to direct synthesis. In the clone expressing Fuc-TIII and beta3Gal-T5 (CHO-FT-T5), sialyl-Lewis a synthesis is strongly inhibited by swainsonine but not by benzyl-alpha-GalNAc, and sialyl-Lewis x is absent, although it is detected in the clones expressing Fuc-TIII and beta3Gal-T1 (CHO-FT-T1) or Fuc-TIII and beta3Gal-T2 (CHO-FT-T2). Endo-beta-galactosidase treatment of N- glycans prepared from clone CHO-FT-T5 releases (+/-NeuAcalpha2-->3)Galbeta1-->3[Fucalpha1-->4]GlcNAcbeta1-->3Gal but not GlcNAcbeta1-->3Gal or type 2 chain oligosaccharides, which are found in CHO-FT-T1 cells. This result indicates that beta3Gal-T5 expression prevents poly-N-acetyllactosamine and sialyl-Lewis x synthesis on N-glycans. Kinetic studies confirm that beta3Gal-T5 prefers acceptors having the GlcNAcbeta1-->3Gal end, including lactotriosylceramide. Competitive reverse transcriptase mediated-polymerase chain reaction shows that the beta3Gal-T5 transcript is expressed in normal colon mucosa but not or poorly in adenocarcinomas. Moreover, recombinant carcinoembryonic antigen purified from a CHO clone expressing Fuc-TIII and beta3Gal-T5 reacts with anti-sialyl-Lewis a and carries type 1 chains on oligosaccharides released by endo-beta-galactosidase. We conclude that beta3Gal-T5 down-regulation plays a relevant role in determining the cancer-associated glycosylation pattern of N-glycans.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenocarcinoma / enzymology*
  • Adenocarcinoma / genetics
  • Adenocarcinoma / metabolism
  • Aged
  • Animals
  • CHO Cells
  • Carcinoembryonic Antigen / metabolism*
  • Colonic Neoplasms / enzymology*
  • Colonic Neoplasms / genetics
  • Colonic Neoplasms / metabolism
  • Cricetinae
  • Down-Regulation
  • Female
  • Fucosyltransferases / biosynthesis
  • Galactosyltransferases / genetics
  • Galactosyltransferases / metabolism*
  • Gastric Mucosa / metabolism
  • Glycoproteins / metabolism*
  • Glycosylation
  • Humans
  • Lewis Blood Group Antigens / metabolism
  • Male
  • Middle Aged
  • Oligosaccharides / metabolism
  • Polysaccharides / metabolism
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sialyl Lewis X Antigen
  • Transfection

Substances

  • Carcinoembryonic Antigen
  • Glycoproteins
  • Lewis Blood Group Antigens
  • Oligosaccharides
  • Polysaccharides
  • Sialyl Lewis X Antigen
  • Fucosyltransferases
  • Galactosyltransferases
  • UDP-galactose N-acetylglucosaminyl-1-3-N-acetylgalactosamine beta-1,3-galactosyltransferase
  • 3-galactosyl-N-acetylglucosaminide 4-alpha-L-fucosyltransferase