trisialoganglioside-gt1 and Carcinoma--Squamous-Cell

The interaction of the urokinase-type plasminogen activator (uPA) receptor (uPAR) with integrins plays a critical role in the regulation of cell adhesion and migration. However, the molecular events underlying the modulation of the interaction of uPAR and integrin are poorly understood. Gangliosides are thought to regulate epithelial cell adhesion and migration by inhibiting alpha(5)beta(1) integrin and epidermal growth factor receptor (EGFR) signaling. We report here that increases in the expression of ganglioside NeuAcalpha2-->3Galbeta1-->3GalNAcbeta1-->4(NeuAcalpha2-->8NeuAcalpha2-->3)Galbeta1-->4Glcbeta1-Cer (GT1b) or NeuAcalpha2-->3Galbeta1-->4Glcbeta1-Cer (GM3) inhibit uPA-dependent cell migration by preventing the association of uPAR with alpha(5)beta(1) integrin or uPAR/alpha(5)beta(1) integrin with the EGFR, respectively. As a result, uPA-dependent focal adhesion kinase (FAK) and integrin-mediated EGFR signaling are suppressed. Both gangliosides inhibit uPAR signaling-stimulated migration; however, GM3 inhibits uPA-induced EGFR phosphorylation by blocking the crosstalk between integrin and EGFR, whereas GT1b suppresses both uPA-induced FAK and EGFR activation by preventing the activation of integrin alpha(5)beta(1).

Topics: Carcinoma, Squamous Cell; Cell Line; Cell Movement; ErbB Receptors; Focal Adhesion Kinase 1; Focal Adhesion Protein-Tyrosine Kinases; G(M3) Ganglioside; Gangliosides; Humans; Integrin alpha5beta1; Keratinocytes; Oligodeoxyribonucleotides, Antisense; Phosphorylation; Protein-Tyrosine Kinases; Receptors, Cell Surface; Receptors, Urokinase Plasminogen Activator; Skin Neoplasms

Polysialoganglioside GT1b, a keratinocyte membrane glycosphingolipid, inhibits normal keratinocyte adhesion and migration on a fibronectin matrix. The specificity of the inhibition for cells plated on a fibronectin matrix and competition of GT1b inhibition with peptide RGDS suggest that GT1b abrogates the alpha 5 beta 1/fibronectin interaction. We examined the effects of GT1b on the adhesion and migration of keratinocyte-derived cell lines and correlated GT1b responsiveness and alpha 5 beta 1 integrin expression. GT1b (5 nM) significantly inhibited migration of normal human keratinocytes, immortalized keratinocytes, and squamous cell carcinoma SCC12F2 cells on fibronectin, but not on collagen I. Concentrations as high as 5 microM had no effect on SCC13 or HaCaT cells. Likewise, GT1b inhibited fibronectin-dependent cell adhesion of normal human keratinocytes, immortalized keratinocytes, and SCC12F2 cells, but had no effect on SCC13 or HaCaT cells. Flow cytometric and Western immunoblot analysis of integrin expression showed significantly decreased alpha 5 and beta 1 integrin expression in SCC13 and HaCaT cells compared to normal keratinocytes, immortalized keratinocytes, and SCC12F2 cells. Incubation with TGF-beta 1 increased alpha 5 beta 1 integrin expression and induced responsiveness to GT1b in HaCaT cells. These data imply that GT1b "response" requires sufficient expression of alpha 5 beta 1 and further suggest that the mechanism of the inhibitory effect of GT1b involves GT1b/alpha 5 beta 1 interaction.

Topics: Carcinoma, Squamous Cell; Cell Adhesion; Cell Line, Transformed; Cell Movement; Cell Transformation, Viral; Cells, Cultured; Depression, Chemical; Facial Neoplasms; Fibronectins; Flow Cytometry; Gangliosides; Humans; Keratinocytes; Male; Neoplasm Proteins; Receptors, Fibronectin; Transforming Growth Factor beta; Tumor Cells, Cultured