The enzyme β1,4-N-acetylglucosaminyltransferase III (GnT-III) catalyzes the addition of a bisecting GlcNAc residue to glycoproteins, resulting in a modulation in biological function. Our previous studies showed that the transfection of the GnT-III gene into B16 melanoma cells results in a suppression of invasive ability and lung colonization. The suppression has been postulated to be due to an increased level of E-cadherin expression on the cell surface, which in turn leads to the up-regulation of cell-cell adhesion. In this study, we report on the effects of overexpression of GnT-III on cell-matrix adhesion. The overexpression of GnT-III, but not that of an enzymatic inactive GnT-III (D323A), inhibits cell spreading and migration on fibronectin, a specific ligand for integrin α5β1, and the focal adhesion kinase phosphorylation. E4-PHA lectin blot analyses showed that the levels of bisecting GlcNAc structures on the integrin α5 subunit as well as α2 and α3 subunits immunoprecipitated from GnT-III transfectants were substantially increased. In addition, the affinity of the binding of integrin α5β1 to fibronectin was significantly reduced by the introduction of the bisecting GlcNAc, to the α5 subunit. These findings suggest that the modification of N-glycan of integrin by GnT-III inhibits its ligand binding ability, subsequently leading to the down-regulation of integrin-mediated signaling.
This work was partly supported by the 21st Century Center for Excellence program from the Ministry of Education, Culture, Sports, Science and Technology of Japan. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
