␣-Dystroglycan (␣-DG) represents a highly glycosylated cell surface molecule that is expressed in the epithelial cell-basement membrane (BM) interface and plays an essential role in epithelium development and tissue organization. The ␣-DG-mediated epithelial cell-BM interaction is often impaired in invasive carcinomas, yet roles and underlying mechanisms of such an impaired interaction in tumor progression remain unclear. We report here a suppressor function of laminin-binding glycans on ␣-DG in tumor progression. In aggressive prostate and breast carcinoma cell lines, lamininbinding glycans are dramatically decreased, although the amount of ␣-DG and ␤-dystroglycan is maintained. The decrease of lamininbinding glycans and consequent increased cell migration were associated with the decreased expression of ␤3-N-acetylglucosaminyltransferase-1 (␤3GnT1). Forced expression of ␤3GnT1 in aggressive cancer cells restored the laminin-binding glycans and decreased tumor formation. ␤3GnT1 was found to be required for laminin-binding glycan synthesis through formation of a complex with LARGE, thus regulating the function of LARGE. Interaction of the laminin-binding glycans with laminin and other adhesive molecules in BM attenuates tumor cell migratory potential by antagonizing ERK/AKT phosphorylation induced by the components in the ECM. These results identify a previously undescribed role of carbohydrate-dependent cell-BM interaction in tumor suppression and its control by ␤3GnT1 and LARGE.glycosylation ͉ cell adhesion ͉ basement membrane ͉ carcinoma I nteraction of epithelial cells with basement membrane (BM) is mediated by cell adhesion molecules, which operate at the interface of epithelial cell-ECM and regulate cell growth, motility, and differentiation by integrating signals from ECM or soluble factors (1-3). One of the most important epithelial cell-BM interactions is mediated by ␣-dystroglycan (␣-DG) on epithelial cells (4).␣-DG is a cell surface receptor for several major BM proteins, including laminin, perlecan, and agrin. A laminin G-like domain in all these glycoproteins binds to a unique glycan structure attached to ␣-DG, and this interaction has been shown to be critical in assembling BM (5, 6). This unique glycan structure is referred to as laminin-binding glycans hereafter. ␣-DG is not attached directly to the plasma membrane but is bound to it through attachment to the transmembrane protein ␤-dystroglycan (␤-DG), which binds to the cytoplasmic protein dystrophin, which, in turn, binds to the actin cytoskeleton and many adaptor molecules involved in cellular signaling (4,5).␣-DG is highly glycosylated and contains both N-linked glycans and mucin type O-glycans. The mucin type O-glycans are clustered in a mucin-like domain at the N-terminal of mature ␣-DG, which includes unique O-mannosyl glycans and sialic acid ␣233Gal␤134GlcNAc␤132Man␣13Ser/Thr (7). Defects in glycosylation of the O-mannosyl glycans have been shown to cause muscular dystrophy (8). So far, 7 glycosyltransferases or glycosyltransferase-like ...

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PST and STX are polysialyltransferases that form polysialic acid in the neural cell adhesion molecule (NCAM), and these two polysialyltransferases often exist together in the same tissues. To determine the individual and combined roles of PST and STX in polysialic acid synthesis, in the present study we asked if PST and STX differ in the acceptor requirement and if PST and STX act together in polysialylation of NCAM. We first examined whether PST and STX differ in the requirement of sialic acid and core structures of N-glycans attached to NCAM. Polysialic acid was formed well on Lec4 and Lec13 cells, which are defective in N-acetylglucosaminyltransferase V and GDP-fucose synthesis, respectively, demonstrating that a side chain elongating from GlcNAc␤136Man␣136R and ␣-1,6-linked fucose are not required. PST and STX were found to add polysialic acid on NCAM⅐Fc molecules sialylated by ␣-2,3-or ␣-2,6-linkage in vitro, but not on NCAM⅐Fc lacking either sialic acid. These results indicate that both PST and STX have relatively broad specificity on N-glycan core structures in NCAM and no remarkable difference exists between PST and STX for the requirement of core structures and sialic acid attached to the N-glycans of NCAM. We then, using various N-glycosylation site mutants of NCAM, discovered that PST strongly prefer the sixth N-glycosylation site, which is the closest to the transmembrane domain, over the fifth site. STX slightly prefer the sixth N-glycosylation site over the fifth N-glycosylation site. The results also demonstrated that polysialic acid synthesized by PST is larger than that synthesized by STX in vitro. Moreover, a mixture of PST and STX more efficiently synthesized polysialic acid on NCAM than PST or STX alone. These results suggest that polysialylation of NCAM is influenced by the difference between PST and STX in their preference for N-glycosylation sites on NCAM. The results also suggest that PST and STX form polysialylated NCAM in a synergistic manner.Polysialic acid is a developmentally regulated carbohydrate composed of a linear homopolymer of ␣-2,8-linked sialic acid (1). NCAM 1 is highly polysialylated in embryonic tissues. In contrast, the majority of NCAM in adult tissues lacks this unique glycan, but polysialylated NCAM is present in the olfactory bulb and hippocampus of adult brain, where neural regeneration persists (2, 3). There is increasing evidence that polysialylated NCAM promotes cell migration and enhances neurite outgrowth and branching during development and neural regeneration (2-4). Polysialic acid is thought to modulate the functional properties of NCAM by rendering it less adhesive to itself (homophilic binding) (5, 6) or to other cell surface molecules (heterophilic binding). In the latter case, it has been shown that NCAM engages in interactions with L1 on the same membrane (cis-interaction) (7).The studies on NCAM knockout mice demonstrated an abnormal formation in the olfactory bulb and hippocampus and a defect in spatial learning and memory (8, 9). By using NCAM knocko...

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