The Two Endocytic Pathways Mediated by the Carbohydrate Recognition Domain and Regulated by the Collagen-like Domain of Galectin-3 in Vascular Endothelial Cells

Abstract

Galectin-3 plays an important role in endothelial morphogenesis and angiogenesis. We investigated the endocytosis of galectin-3 in human vascular endothelial cells and showed that galectin-3 could associate with and internalized into the cells in a carbohydrate-dependent manner. Our work also revealed that galectin-3 was transported to the early/recycling endosomes and then partitioned into two routes – recycling back to the plasma membrane or targeting to the late endosomes/lysosomes. Various N- and C-terminal truncated forms of galectin-3 were constructed and compared with the full-length protein. These comparisons showed that the carbohydrate-recognition domain of galectin-3 was required for galectin-3 binding and endocytosis. The N-terminal half of the protein, which comprises the N-terminal leader domain and the collagen-like internal repeating domain, could not mediate binding and endocytosis alone. The collagen-like domain, although it was largely irrelevant to galectin-3 trafficking to the early/recycling endosomes, was required for targeting galectin-3 to the late endosomes/lysosomes. In contrast, the leader domain was irrelevant to both binding and intracellular trafficking. The data presented in this study correlate well with different cellular behaviors induced by the full-length and the truncated galectin-3 and provide an alternative way of understanding its angiogenic mechanisms.

Galectin-3 Guides Intracellular Trafficking of Some Human Serotransferrin Glycoforms

“…Galectin-3 has recently been shown to be rapidly internalized by vascular endothelial cells (40) and macrophages (39) appearing in EEA1 positive endosomes within 10 -15 min and then partitioned into two routes, either recycling (through Rab11 positive endosomes) or targeting to the LE/lysosomes (LAMP-1 positive endosomes). However, the dynamics of these endocytic pathways partly differed depending on cell type.…”

Multiple approaches to assess pectin binding to galectin-3

“…Potato-galactan (p-galactan) and endo- Recombinant human Gal-3, His-tagged Gal-3, and biotin-labeled Gal-3 were prepared as reported [32,33]. Other reagents were of analytical grade.…”

Building endocytic pits without clathrin

“…Galectin 3 was previously reported to have a stimulatory or an inhibitory effect on endocytosis [86][87][88][89] . The finding that CIE of the CLIC cargo CD44 was stimulated at con centrations of galectin 3 between 0.01 μg ml −1 and 1 μg ml −1 but inhibited at 10 μg ml −1 (REF.…”