Abstract
Abstract: :
Purpose: We have previously shown that carbohydrate–binding proteins, galectins–3 and –7, are expressed in corneal epithelium and that the exogenous addition of these galectins stimulate the re–epithelialization of corneal wounds in a mouse animal model (J. Biol. Chem. 277:42299–42305, 2002). Since, most of the extracelluar matrix molecules, growth factors and growth factor receptors are glycosylated, it is logical to hypothesize that one or more of these molecules serve as counterreceptors of galectins and, that the galectins influence re–epithelialization of corneal wound by binding to and influencing the function of the specific counterreceptors. The goal of the present study was to identify the counterreceptors of galectins–3 and –7 and to determine whether the two galectins bind to similar or distinct counterreceptors. Methods: Primary cultures of corneal epithelium were prepared using corneas of 6– to 12–month old mice (C57BL/6 and 129 mixed genetic background). Membrane extracts of corneal epithelial cell cultures were electrophoresed on SDS–polyacrylamide gels and protein blots were probed with biotinylated galectins–3– and –7 in the presence and absence of ß–lactose. In some experiments, the membrane extracts were chromatograhed on galectin affinity columns, the bound material was eluted by ß–lactose and analyzed by lectin blot analysis using biotinylated galectins. Results: Galectin–3 and –7 bound to distinct glycoprotein counterreceptors. Galectin–3 bound to two major (116–kD and 120–kD) and two minor components (74–kD and 64–kD). In contrast, galectin–7 bound to four major (60–kD, 52–kD, 47–kD and 35–kD) and a number of minor components. In each case, binding of the galectins to the glycoproteins was abolished by ß–lactose. Conclusions: Galectins–3 and –7 bind to distinct counterreceptors. Studies are underway to establish the identity of these receptors. This study should help understand the molecular mechanism by which galectins influence re–epithelialization of corneal wounds.
Keywords: cell adhesions/cell junctions • glycoconjugates/glycoproteins • cornea: epithelium