Contributions of Galectin-3 and -9 to Epithelial Cell Adhesion Analyzed by Single Cell Force Spectroscopy

Jens Friedrichs,Juha M Torkko,Jonne Helenius,Terhi P Teräväinen,Joachim Füllekrug,Daniel J Muller,Kai Simons,Aki Manninen

doi:10.1074/jbc.m701867200

Abstract

Galectins are widely expressed in epithelial tissues and have been implicated in a variety of cellular processes, including adhesion and polarization. Here we studied the contributions of galectins in cell adhesion and cyst formation of Madin-Darby canine kidney cells. Quantitative single cell force spectroscopy and standard adhesion assays were employed to study both early (<2 min) and long term (90 min) adhesion of cells to different extracellular matrix components. Inhibitors were used to examine the contribution of integrins and galectins in general and RNA interference to specifically address the role of two abundantly expressed galectins, galectin-3 and -9. We found that both galectin-3 and -9 were required for optimal long term cell adhesion to both collagen I and laminin-111. Early adhesion to laminin was found to be integrin-independent and was instead mediated by carbohydrate interactions and galectin-3 and -9. The opposite was observed for early adhesion to collagen. Although similar, the contributions of galectin-3 and -9 to adhesion appeared to be by distinct processes. These defects in adhesion of the two galectin knockdown cell lines may underlie the epithelial phenotypes observed in the cyst assays. Our findings emphasize the complex regulation of epithelial cell functions by galectins.

Highlights

Galectins constitute a family of structurally related proteins that bind to ␤-galactoside residues and interact with a broad spectrum of ligands [5, 6]
We studied the expression of several galectin family members in Madin-Darby canine kidney (MDCK) cells and used RNA interference (RNAi) to address the role of abundantly expressed galectin-3 and galectin-9 in cell adhesion and epithelial morphogenesis
Galectin-3 was expressed at very high levels

Summary

Introduction

Galectins constitute a family of structurally related proteins that bind to ␤-galactoside residues and interact with a broad spectrum of ligands [5, 6]. Galectin-9 Secretion—MDCK-Gal9myc cells grown for 3– 4 days on 12-mm Transwell filters were washed twice with starvation medium (EMEM lacking methionine and cysteine (PAA Laboratories), containing L-glutamine and antibiotics) before a 15-min incubation in starvation medium at 37 °C.

Results

Conclusion