Intracellular sorting of galectin-8 based on carbohydrate fine specificity.
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.
Carlsson
Susanne
S
Institute of Laboratory Medicine, Section of Microbiology Immunology and Glycobiology, Lund University, Sölvegatan 23, S-223 62 Lund, Sweden. susanne.carlsson@med.lu.se
Carlsson
Michael C
MC
Leffler
Hakon
H
D000584:Amiloride / Q000737:chemistry; D000818:Animals; D016466:CHO Cells; D002241:Carbohydrates / Q000737:chemistry; D002462:Cell Membrane / Q000378:metabolism; D002746:Chlorpromazine / Q000494:pharmacology; D006224:Cricetinae; D003412:Cricetulus; D015638:Cytochalasin D / Q000494:pharmacology; D004705:Endocytosis; D037161:Galectins / Q000737:chemistry / Q000502:physiology*; D018922:HL-60 Cells; D006801:Humans; D007700:Kinetics; D012794:Sialic Acids / Q000737:chemistry; D020298:U937 Cells; D047392:beta-Cyclodextrins / Q000737:chemistry
D002241:Carbohydrates; D037161:Galectins; C091991:LGALS8 protein, human; D012794:Sialic Acids; D047392:beta-Cyclodextrins; C108732:methyl-beta-cyclodextrin; D015638:Cytochalasin D; D000584:Amiloride; D002746:Chlorpromazine