Suppression of lung metastasis of B16 mouse melanoma by N-acetylglucosaminyltransferase III gene transfection.

Abstract

The j31-6 structure of N-linked oligosaccharides, formed by j8-1,6-N-acetylglucosaminyltransferase (GnT-V), is associated with metastatic potential. We established a highly metastatic subclone, B16-hm, from low metastatic B16-F1 murine melanoma cells. The gene encoding 18-1,4-N-acetylglucosaminyltransferase (GnT-III) was introduced into the B16-hm cells, and three clones that stably expressed high GnT-III activity were obtained. In these transfectants, the affinity to leukoagglutinating phytohemagglutinin was reduced, whereas the binding to erythroagglutinating phytohemagglutinin was increased, indicating that the level of 31-6 structure was decreased due to competition for substrate between intrinsic GnT-V and ectopically expressed GnT-III.Lung metastasis after intravenous injection of the transfectants into syngeneic and nude mice was significantly suppressed, suggesting that the decrease in 81-6 structure suppressed metastasis via a mechanism independent of the murine system. These transfectants also displayed decreased invasiveness into Matrigel and inhibited cell attachment to collagen and laminin. Cell growth was not affected. Our results demonstrate a causative role for p1-6 branches in invasion and cell attachment in the extravasation stage of metastasis.Malignant transformation is highly associated with alterations of N-linked oligosaccharides. The extensive and persistent changes in the population of protein-bound oligosaccharides in malignant cells have been proposed to disturb cell division, decrease intercellular adhesiveness, and mask immunogenicity (1). The glycopeptides in polyoma-transformed baby hamster kidney cells have unusually high molecular weights due to the presence of external sialic acid as well as galactose and GlcNAc (2). The predominant surface glycopeptide of baby hamster kidney cells transformed by Rous sarcoma virus was determined to be a triantennary, completely sialylated, complex glycopeptide containing a core region of Man, GlcNAc, and Fuc (3). Synthesis of these elongated branches is initiated by 13-1,6-N-acetylglucosaminyltransferase (GnT-V; EC 2.4.1.155), which catalyzes the formation of the (31-6 branch (4). GnT-V activity correlated well with the metastatic potential of rastransformed Rat2 fibroblasts, SP1 mammary carcinoma cells, the MDAY-D2 lymphoma cell line (5, 6), and human colon cancer cells (7). Rat2 fibroblasts transfected with H-ras or v-frs exhibited metastatic potential and had elevated GnT-V activity and increased (1-6 branches (8), whereas a mutant with decreased GnT-V activity from a highly metastatic tumor cell line had a decreased potential for metastasis in mice (6). These observations have suggested a positive correlation between ,B1-6 branching and metastatic capacity.One approach to analyzing the role of GnT-V and its product, (31-6 branches, in metastasis is to reduce GnT-V activity in malignant cells with a high tendency to metastasize. As shown in Fig. 1, both 1-1,4-N-acetylglucosaminyltransferase (GnT-III; EC 2.4.1.144) and GnT-V use the t...

Potential roles of N-glycosylation in cell adhesion

“…Overexpression of GnT-III partially inhibited EMT induced by TGF-β1 [61]. Similar to previous studies [53,54], GnT-III modified E-cadherin, which served to prolong E-cadherin turnover on the cell surface. Furthermore, GnT-III expression consistently inhibited β-catenin translocation from cell-cell contact into the cytoplasm and nucleus.…”

“…In an earlier study, Yoshimura, et al reported that E-cadherin-mediated cell adhesion was regulated by GnT-III. Overexpression of GnT-III increased the retention of Ecadherin at the cell border, which might result in an enhancement of E-cadherin-mediated homotypic adhesion [53,54]. Thus, expression of E-cadherin may be regulated, not only by transcriptional factors, but also by posttranscriptional processing, maturation and modifications.…”

Deficiency in surface expression of E‐selectin ligand promotes lung colonization in a mouse model of breast cancer

“…20 A phenotype similar to what we observed has been associated with an increase in b-1fi6-GlcNAc branching structures on complex N-linked carbohydrates. 51 Replacing b1fi6 branching structures with bisecting GlcNAc has led to enhanced homotypic adhesion of tumor cells via increased functional expression of E-cadherin. 52 It has also been suggested that integrins are involved in lung metastasis and that the tumor a 3 b 1 FIGURE 4 -sLe x -negative cells metastasize to the lung more than sLe x -positive and original 4T1 cells.…”

Lentiviral Vector-Mediated shRNA against AIMP2-DX2 Suppresses Lung Cancer Cell Growth through Blocking Glucose Uptake

“…Gene expression of GnT-V is regulated by a transcriptional factor involved in angiogenesis and invasion of tumor cells (Taniguchi et al, 1999). When the formation of the product of GnT-V is inhibited by overexpression of GnT-III, lung metastasis of melanoma cells is suppressed (Yoshimura et al, 1995). GnT-III catalyzes the addition of bisecting GlcNAc (β-1,4-GlcNAc branching) to biantennary sugar chains and prevents the subsequent actions of GnT-V and GnT-IV (Sasai et al, 2002;Sultan et al, 1997;Wang et al, 1997).…”