Intracellular sorting of galectin-8 based on carbohydrate fine specificity

Abstract

Galectin-8 has two carbohydrate recognition domains (CRDs), both of which bind beta-galactosides, but have different fine specificity for larger saccharides. Previously we found that both CRDs were needed for efficient cell surface binding and signaling by soluble galectin-8, but unexpectedly binding of the N-CRD to its best ligands, alpha2-3-sialylated galactosides, was not needed. In search for another role for this fine specificity, we now compared endocytosis of galectin-8 in Chinese hamster ovary (CHO) cells and in a mutant (Lec2) lacking sialylated glycans, by fluorescence microscopy. Galectin-8 was endocytosed in both cells by a non-clathrin and non-cholesterol dependent pathway, but surprisingly, the pathway after endocytosis differed dramatically. In wild type (wt) cells, galectin-8 was found along the plasma membrane, near the nucleus, and in small vesicles. In the Lec2 cells, galectin-8 was found in larger vesicles evenly spread in the cell, but not along the plasma membrane or near the nucleus. A galectin-8 mutant with an N-CRD having reduced affinity to sialylated glycans and increased affinity for other glycans, gave a Lec2 like pattern in the wt CHO cells, but a wt pattern in the Lec2 cells. Moreover, the pattern of galectin-3 after endocytosis differed from that of both the wt and mutant galectin-8. These data clearly demonstrate a role of galectin fine specificity for intracellular targeting.

Dimeric Galectin-8 Induces Phosphatidylserine Exposure in Leukocytes through Polylactosamine Recognition by the C-terminal Domain

“…Consistent with this, a recent study on the N-terminal domain of Gal-9 (Gal-9N) demonstrated that it is also a dimer, as evidenced by studies of the crystal structure and solution-based experiments (53). Although we found that only the C-terminal domain of Gal-8 binds the functional signaling receptors on HL60 cells, the unique binding properties of each separate domain corroborates previously studies (29,35,54) and suggests that different cells may possess unique sensitivity to the potential signaling effects of each individual domain (28,55). Furthermore, because the N-terminal domain of Gal-8 intrinsically dimerizes, cleavage of the linker region between Gal-8N and Gal-8C may allow regulatory circuits to dissect potential signaling pathways initiated by each separate domain.…”

Galectin-3 Guides Intracellular Trafficking of Some Human Serotransferrin Glycoforms

“…Galectin-3 was rapidly taken up by the SKBR3 cells, as expected from other cell types (33,34) (Fig. 4B), and was after 24 h found in both large and small vesicular structures in the cytosol (Fig.…”

Galectin-1 and Galectin-8 Have Redundant Roles in Promoting Plasma Cell Formation

“…Notably, the N-terminal CRD of Gal-8 can bind to type 1 LacNAc, but the C-terminal CRD of Gal-8 has a higher binding affinity for poly-LacNAc structures (40). The differential biological activities of the N-versus C-terminal CRD of Gal-8 have been shown (41), including in killing E. coli strains (42), in activating platelets (43), and in modulating neutrophil function (44); however, which CRD of Gal-8 is responsible for its positive role in plasma cell formation remains to be determined. In the current study, sulfomodified Gals or GlcNAc on type 1 or type 2 LacNAcs were used as competitors to elucidate the binding ligands for Gal-1 and Gal-8 on mature B cell surfaces.…”