Differential effect of GalNAcalpha-O-bn on intracellular trafficking in enterocytic HT-29 and Caco-2 cells: correlation with the glycosyltransferase expression pattern.
Our previous work has shown that long-term treatment of mucus-secreting HT-29 cells with 1-benzyl-2-acetamido-2-deoxy-alpha-D-galactopyranoside (GalNAcalpha-O-bn), a competitive inhibitor of O-glycosylation, induced several phenotypic changes, in particular a blockade in the secretion of mucins, which are extensively O-glycosylated glycoproteins. Here, we have analyzed the effects of GalNAcalpha-O-bn upon the intracellular trafficking of basolateral and apical membrane glycoproteins at the cellular and biochemical levels in two types of cells, HT-29 G(-) and Caco-2, differentiated into an enterocyte-like phenotype. In HT-29 G(-) cells, but not in Caco-2 cells, DPP-IV and CD44 failed to be targeted to the apical or basolateral membrane, respectively, and accumulated inside intracytoplasmic vesicles together with GalNAcalpha-O-bn metabolites. We observed a strong inhibition of alpha2,3-sialylation of glycoproteins in HT-29 G(-) cells correlated to the high expression of alpha2,3-sialyltransferases ST3Gal I and ST3Gal IV. In these cells, DPP-IV and CD44 lost the sialic acid residue substituting the O-linked core 1 structure Galbeta1-3GalNAc (T-antigen). In contrast, sialylation was not modified in Caco-2 cells, but a decrease of alpha1,2-fucosylation was observed, in correlation with the high expression of alpha1,2-fucosyltransferases Fuc-TI and Fuc-TII. In conclusion, in HT-29 G(-) cells, GalNAcalpha-O-bn induces a specific cellular phenotype, which is morphologically characterized by the formation of numerous intracellular vesicles, in which are accumulated defectively sialylated O-glycosylproteins originally targeted to basolateral or apical membranes, and GalNAcalpha-O-bn metabolites.
Gouyer
V
V
Unité INSERM 377, place de Verdun, France.
Leteurtre
E
E
Delmotte
P
P
Steelant
W F
WF
Krzewinski-Recchi
M A
MA
Zanetta
J P
JP
Lesuffleur
T
T
Trugnan
G
G
Delannoy
P
P
Huet
G
G
D018938:Caco-2 Cells / Q000378:metabolism; D002454:Cell Differentiation; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases / Q000187:drug effects / Q000378:metabolism; D004789:Enzyme Activation / Q000502:physiology; D000939:Epitopes / Q000276:immunology / Q000378:metabolism; D005647:Fucosyltransferases / Q000235:genetics* / Q000378:metabolism; D005690:Galactose / Q000008:administration & dosage* / Q000031:analogs & derivatives* / Q000378:metabolism*; D015870:Gene Expression / Q000235:genetics; D006031:Glycosylation / Q000187:drug effects*; D019073:HT29 Cells / Q000378:metabolism; D006801:Humans; D018960:Hyaluronan Receptors / Q000187:drug effects / Q000378:metabolism; D008562:Membrane Glycoproteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D009005:Monosaccharides / Q000737:chemistry / Q000378:metabolism; D011134:Polysaccharides / Q000276:immunology / Q000378:metabolism; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology*; D012799:Sialyltransferases / Q000235:genetics* / Q000378:metabolism; D014407:Tumor Cells, Cultured
C411946:1-benzyl-2-acetamido-2-galactopyranoside; D000939:Epitopes; D018960:Hyaluronan Receptors; D008562:Membrane Glycoproteins; D009005:Monosaccharides; D011134:Polysaccharides; D005647:Fucosyltransferases; D012799:Sialyltransferases; C415802:DP IV-related protease, human; D004152:Dipeptidyl-Peptidases and Tripeptidyl-Peptidases; D005690:Galactose