GalNAc-alpha-O-benzyl inhibits NeuAcalpha2-3 glycosylation and blocks the intracellular transport of apical glycoproteins and mucus in differentiated HT-29 cells.
Exposure for 24 h of mucus-secreting HT-29 cells to the sugar analogue GalNAc-alpha-O-benzyl results in inhibition of Galbeta1-3GalNAc:alpha2,3-sialyltransferase, reduced mucin sialylation, and inhibition of their secretion (Huet, G., I. Kim, C. de Bolos, J.M. Loguidice, O. Moreau, B. Hémon, C. Richet, P. Delannoy, F.X. Real., and P. Degand. 1995. J. Cell Sci. 108:1275-1285). To determine the effects of prolonged inhibition of sialylation, differentiated HT-29 populations were grown under permanent exposure to GalNAc-alpha-O-benzyl. This results in not only inhibition of mucus secretion, but also in a dramatic swelling of the cells and the accumulation in intracytoplasmic vesicles of brush border-associated glycoproteins like dipeptidylpeptidase-IV, the mucin-like glycoprotein MUC1, and carcinoembryonic antigen which are no longer expressed at the apical membrane. The block occurs beyond the cis-Golgi as substantiated by endoglycosidase treatment and biosynthesis analysis. In contrast, the polarized expression of the basolateral glycoprotein GP 120 is not modified. Underlying these effects we found that (a) like in mucins, NeuAcalpha2-3Gal-R is expressed in the terminal position of the oligosaccharide species associated with the apical, but not the basolateral glycoproteins of the cells, and (b) treatment with GalNAc-alpha-O-benzyl results in an impairment of their sialylation. These effects are reversible upon removal of the drug. It is suggested that alpha2-3 sialylation is involved in apical targeting of brush border membrane glycoproteins and mucus secretion in HT-29 cells.
Huet
G
G
Unité de Recherches sur la Biologie et la Physiopathologie des Cellules Mucipares, Institut National de la Sante et de la Recherche Medicale (INSERM) U377, 59045 Lille Cedex, France.
Hennebicq-Reig
S
S
de Bolos
C
C
Ulloa
F
F
Lesuffleur
T
T
Barbat
A
A
Carrière
V
V
Kim
I
I
Real
F X
FX
Delannoy
P
P
Zweibaum
A
A
D000116:Acetylgalactosamine / Q000031:analogs & derivatives* / Q000494:pharmacology; D001593:Benzyl Compounds / Q000494:pharmacology*; D001692:Biological Transport; D002454:Cell Differentiation; D004305:Dose-Response Relationship, Drug; D006023:Glycoproteins / Q000378:metabolism; D006031:Glycosylation / Q000187:drug effects; D006056:Golgi Apparatus / Q000378:metabolism; D019073:HT29 Cells; D006801:Humans; D008871:Microvilli / Q000378:metabolism; D009077:Mucins / Q000378:metabolism; D009093:Mucus; D019158:N-Acetylneuraminic Acid / Q000037:antagonists & inhibitors*; D009438:Neuraminic Acids / Q000378:metabolism; D009844:Oligosaccharides / Q000378:metabolism
D001593:Benzyl Compounds; D006023:Glycoproteins; D009077:Mucins; D009438:Neuraminic Acids; D009844:Oligosaccharides; C032592:N-glycolylneuraminic acid; C080739:benzyl-alpha-N-acetylgalactosamine; D019158:N-Acetylneuraminic Acid; D000116:Acetylgalactosamine