Abstract
PURPOSE: To study the relation of N-glycan structure on cell surface glycoproteins to the metastatic phenotypes. METHODS: Two human hepatocarcinoma 7721 cell lines transfected with sense or antisense cDNA of GnT-V, named GnT-V/7721 and GnT-V-AS/7721, respectively, were adopted, because GnT-V can change the antennary number and the content of the beta 1,6 GlcNAc branch in N-glycans. The effects of over- and under-expression of GnT-V on the metastasis-associated phenotype of the transfected cells were investigated and compared with the cells mock-transfected with the plasmid vector. RESULTS: In GnT-V/7721 cells, GnT-V activity was increased by 92% compared with the mock cells. HRP-labeled lectin staining of transfected cells showed elevated intensity with HRP-L-PHA and reduced intensity with HRP-ConA, suggesting the increased antennary number and content of the beta 1,6 GlcNAc branch in N-glycans. Analysis of the N-glycan structure of [3H]-labeled glycopeptides prepared from cell-surface [3H] glycoproteins using DSA-affinity chromatography also revealed the above change of the N-glycan structure in a more quantitative manner. GnT-V/7721 cells showed a suppressed cell attachment to fibronectin (Fn) or laminin (Ln), and increased cell migration and invasion through matrigel. In contrast, GnT-V-AS/7721 cells showed reduction of both GnT-V activity and content of the beta 1,6 branch in N-linked glycans, elevation of cell attachment to Fn or Ln, and decline of cell migration and invasion through matrigel. These changes were just the opposite to those in GnT-V/7721 cells. CONCLUSIONS: The alteration of N-glycan structure in surface glycoproteins resulting from the activity change of GnT-V contributes to the alterations in metastasis-associated phenotypes. The product of GnT-V, the beta 1,6 GlcNAc branch in N-linked glycans, is a structural factor of adhesion inhibition and invasion promotion. GnT-V is, therefore, closely related to cancer metastasis and its over-expression is an important molecular mechanism of metastasis.