Extended core 1 and core 2 branched O-glycans differentially modulate sialyl Lewis X-type L-selectin ligand activity.
It has been established that sialyl Lewis x in core 2 branched O-glycans serves as an E- and P-selectin ligand. Recently, it was discovered that 6-sulfosialyl Lewis x in extended core 1 O-glycans, NeuNAcalpha2-->3Galbeta1-->4(Fucalpha1-->3(sulfo-->6))GlcNAcbeta1--> 3Galbeta1-->3GalNAcalpha1-->Ser/Thr, functions as an L-selectin ligand in high endothelial venules. Extended core 1 O-glycans can be synthesized when a core 1 extension enzyme is present. In this study, we first show that beta1,3-N-acetylglucosaminyltransferase-3 (beta3GlcNAcT-3) is almost exclusively responsible for core 1 extension among seven different beta3GlcNAcTs and thus acts on core 1 O-glycans attached to PSGL-1. We found that transcripts encoding beta3GlcNAcT-3 were expressed in human neutrophils and lymphocytes but that their levels were lower than those of transcripts encoding core 2 beta1,6-N-acetylglucosaminyltransferase I (Core2GlcNAcT-I). Neutrophils also expressed transcripts encoding fucosyltransferase VII (FucT-VII) and Core2GlcNAcT-I, whereas lymphocytes expressed only small amounts of transcripts encoding FucT-VII. To determine the roles of sialyl Lewis x in extended core 1 O-glycans, Chinese hamster ovary (CHO) cells were stably transfected to express PSGL-1, FucT-VII, and either beta3GlcNAcT-3 or Core2GlcNAcT-I. Glycan structural analyses disclosed that PSGL-1 expressed in these transfected cells carried comparable amounts of sialyl Lewis x in extended core 1 and core 2 branched O-glycans. In a rolling assay, CHO cells expressing sialyl Lewis x in extended core 1 O-glycans supported a significant degree of shear-dependent tethering and rolling of neutrophils and lymphocytes, although less than CHO cells expressing sialyl Lewis x in core 2 branched O-glycans. These results indicate that sialyl Lewis x in extended core 1 O-glycans can function as an L-selectin ligand and is potentially involved in neutrophil adhesion on neutrophils bound to activated endothelial cells.
Mitoma
Junya
J
Glycobiology Program, Cancer Research Center, the Burnham Institute, La Jolla, California 92037, USA.
Petryniak
Bronislawa
B
Hiraoka
Nobuyoshi
N
Yeh
Jiunn-Chern
JC
Lowe
John B
JB
Fukuda
Minoru
M
D000818:Animals; D016466:CHO Cells; D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D005434:Flow Cytometry; D006367:HeLa Cells; D006801:Humans; D019041:L-Selectin / Q000737:chemistry* / Q000378:metabolism; D008024:Ligands; D008214:Lymphocytes / Q000276:immunology / Q000378:metabolism; D008954:Models, Biological; D008969:Molecular Sequence Data; D017351:N-Acetylglucosaminyltransferases / Q000737:chemistry* / Q000276:immunology; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry* / Q000276:immunology; D011134:Polysaccharides / Q000737:chemistry*; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000080506:Sialyl Lewis X Antigen; D014162:Transfection
D008024:Ligands; D009844:Oligosaccharides; D011134:Polysaccharides; D012333:RNA, Messenger; D000080506:Sialyl Lewis X Antigen; D019041:L-Selectin; D017351:N-Acetylglucosaminyltransferases; C428133:beta-1,3-N--acetylglucosaminyltransferase T3