Exploration into Galectin-3 Driven Endocytosis and Lattices

Essentially all plasma membrane proteins are glycosylated, and their activity is regulated by tuning their cell surface dynamics. This is achieved by glycan-binding proteins of the galectin family that either retain glycoproteins within lattices or drive their endocytic uptake via the clathrin-independent glycolipid-lectin (GL-Lect) mechanism. Here, we have used immunofluorescence-based assays to analyze how lattice and GL-Lect mechanisms affect the internalization of the cell adhesion and migration glycoprotein αβ integrin. In retinal pigment epithelial (RPE-1) cells, internalized αβ integrin is found in small peripheral endosomes under unperturbed conditions. Pharmacological compounds were used to competitively inhibit one of the galectin family members, galectin-3 (Gal3), or to inhibit the expression of glycosphingolipids, both of which are the fabric of the GL-Lect mechanism. We found that under acute inhibition conditions, endocytic uptake of αβ integrin was strongly reduced, in agreement with previous studies on the GL-Lect driven internalization of the protein. In contrast, upon prolonged inhibitor treatment, the uptake of αβ integrin was increased, and the protein was now internalized by alternative pathways into large perinuclear endosomes. Our findings suggest that under these prolonged inhibitor treatment conditions, αβ integrin containing galectin lattices are dissociated, leading to an altered endocytic compartmentalization.