Core2 O-Glycan Structure Is Essential for the Cell Surface Expression of Sucrase Isomaltase and Dipeptidyl Peptidase-IV during Intestinal Cell Differentiation

Abstract

Alterations in glycosylation play an important role during intestinal cell differentiation. Here, we compared expression of mucin-type O-glycan synthases from proliferating and differentiated HT-29 and Caco-2 cells. Mucin-type O-glycan structures were analyzed at both stages by mass spectrometry. Core2 ␤1,6-N-acetylglucosaminyltransferase-2 (C2GnT-2) was markedly increased in differentiated HT-29 and Caco-2 cells, but the core3 structure was hardly detectable. To determine whether such differential expression of mucin-type O-glycan structures has physiological significance in intestinal cell differentiation, expression of sucrase isomaltase (SI) and dipeptidylpeptidase IV (DPP-IV), two well known intestinal differentiation markers, was examined. Interestingly, the fully glycosylated mature form of SI was decreased in C2GnT-2 knock-out mice but not in core2 N-acetylglucosaminyltransferase-3 (C2GnT-3) nulls. In addition, expression of SI and DPP-IV was dramatically reduced in C2GnT-1-3 triple knock-out mice. These patterns were confirmed by RNAi analysis; C2GnT-2 knockdown significantly reduced cell surface expression of SI and DPP-IV in Caco-2 cells. Similarly, overexpression of the core3 structure in HT-29 cells attenuated cell surface expression of both enzymes. These findings indicate that core3 O-glycan structure regulates cell surface expression of SI and DPP-IV and that core2 O-glycan is presumably an essential mucin-type O-glycan structure found in both molecules in vivo. Finally, goblet cells in the upper part of the crypt showed impaired maturation in the core2 O-glycan-deficient mice. These studies are the first to clearly identify functional mucin-type O-glycan structures modulating cell surface expression of SI and DPP-IV during the intestinal cell differentiation.

Core-glycosylated Mucin-like Repeats from MUC1 Are an Apical Targeting Signal

“…The polarized distribution of surface 35 S-labeled MUC1-22TR, Tac, and chimeras observed at 120 min of chase (Figs. 2 and 3) also reflects the steady state expression of these proteins when their surface expression is analyzed by confocal immunofluorescence microscopy (Fig.…”

Emerging Paradigms for the Initiation of Mucin-type Protein O-Glycosylation by the Polypeptide GalNAc Transferase Family of Glycosyltransferases

“…x O-linked glycan 5 at a specific N-terminal Thr residue of PSGL1 (Thr-57 in human or Thr-58 in murine PSGL1 (72,73)). The targeted biosynthesis of this O-glycan structure at this site is not understood; however, in the ppGalNAc T1 knock-out mouse PSGL1 is significantly reduced (74).…”

“…Consequently, glycoproteins containing O-glycosylated mucin domains function in the protection of the cell surface, the modulation of cell-cell interactions, in the inflammatory and immune response, in metastasis and tumorigenesis, and in protein sorting, targeting, and turnover (for examples see Refs. [1][2][3][4][5]. It is also likely that such O-glycosylated domains may further present a molecular code for the specific recognition of additional binding partners, enzymes, or even other glycosyltransferases.…”

“…Mucin-type protein O-glycosylation is initiated in the Golgi compartment by the transfer of ␣-GalNAc, from UDPGalNAc, 5 to Ser and Thr residues of polypeptide acceptors by the large family (ϳ20) of UDP-GalNAc:polypeptide ␣-Nacetylgalactosaminyltransferases (ppGalNAc Ts). By subsequent action of a series of specific glycosyltransferases, the O-linked glycan can be further elongated to produce a vast array of glycan structures (14).…”

Core 2 β-1, 6-N-acetylglucosaminyltransferase-1 expression in prostate biopsy specimen is an indicator of prostate cancer aggressiveness

“…Biosynthesis of oligosaccharides on glycoproteins is performed in concert by several glycosyltransferases; GCNT1 is one of the glycosyltransferases that forms the core 2 O-glycans on the surface of lymphocytes and various cancer cells [13,14,26,27]. It is noteworthy that the present study clearly demonstrated that immunohistochemical status of GCNT1 expression on prostate biopsy specimen 11 closely related to extra-prostatic extension, lymph node metastasis of PCa (Table 2).…”