Cell-Cell Interaction-dependent Regulation of N-Acetylglucosaminyltransferase III and the Bisected N-Glycans in GE11 Epithelial Cells: INVOLVEMENT OF E-CADHERIN-MEDIATED CELL ADHESION

Junko Iijima,Yanyang Zhao,Tomoya Isaji,Akihiko Kameyama,Shuuichi Nakaya,Xiangchun Wang,Hideyuki Ihara,Xinyao Cheng,Takatoshi Nakagawa,Eiji Miyoshi,Akihiro Kondo,Hisashi Narimatsu,Naoyuki Taniguchi,Jianguo Gu

doi:10.1074/jbc.m601961200

Abstract

Changes in oligosaccharide structures are associated with numerous physiological and pathological events. In this study, the effects of cell-cell interactions on N-linked oligosaccharides (N-glycans) were investigated in GE11 epithelial cells. N-glycans were purified from whole cell lysates by hydrazinolysis and then detected by high performance liquid chromatography and mass spectrometry. Interestingly, the population of the bisecting GlcNAc-containing N-glycans, the formation of which is catalyzed by N-acetylglucosaminyltransferase III (GnT-III), was substantially increased in cells cultured under dense conditions compared with those cultured under sparse conditions. The expression levels and activities of GnT-III but not other glycosyltransferases, such as GnT-V and alpha1,6-fucosyltransferase, were also consistently increased in these cells. However, this was not observed in mouse embryonic fibroblasts or MDA-MB231 cells, in which E-cadherin is deficient. In contrast, perturbation of E-cadherin-mediated adhesion by treatment with EDTA or a neutralizing anti-E-cadherin antibody abolished the up-regulation of expression of GnT-III. Furthermore, we observed the significant increase in GnT-III activity under dense growth conditions after restoration of the expression of E-cadherin in MDA-MB231 cells. Our data together indicate that a E-cadherin-dependent pathway plays a critical role in regulation of GnT-III expression. Given the importance of GnT-III and the dynamic regulation of cell-cell interaction during tissue development and homeostasis, the changes in GnT-III expression presumably contribute to intracellular signaling transduction during such processes.

Highlights

It has been well known that sugar chains have various effects on the functional aspects of glycoproteins and play important roles in cell dif
In order to investigate the effects of cell-cell interactions on N-glycan biosynthesis, we compared N-glycans obtained from cells cultured under dense or sparse culture conditions
GE11 cells were selected as a model cell to reduce the possible influence of ␤1 integrin-mediated cell adhesion, since GE11 cells are derived from ␤1 integrin knock-out embryonic stem cells [20]

Summary

The abbreviations used are

GnT-V, N-acetylglucosaminyltransferase V; GnT-III, N-acetylglucosaminyltransferase III; Fut8, ␣1,6-fucosyltransferase; PA, pyridylaminated; HPLC, high performance liquid chromatography; MS, mass spectrometry; E4-PHA, erythroagglutinating isolectin; MALDI, matrix-assisted laser desorption ionization; TOF, time-of-flight. Up-regulation of GnT-III by Cell Adhesion in cell recognition, tissue morphogenesis, and tumor suppression [17]. The loss of E-cadherin expression or function in epithelial carcinoma has long been thought to be a primary reason for the disruption of tight epithelial cell-cell contacts and the release of invasive tumor cells from the primary tumor [18]. N-cadherin is found primarily in neural tissues and fibroblasts, where it is thought to mediate a less stable and more dynamic form of cell-cell adhesion [19]. We investigated this issue using a dense or sparse culture model, and found that bisected N-glycans were dramatically induced in cells under conditions of a dense culture through an E-cadherin involved pathway

EXPERIMENTAL PROCEDURES

RESULTS

DISCUSSION