title: Laminin-1 is a novel carrier glycoprotein for the nonsulfated HNK-1 epitope in mouse kidney.
abstract: The HNK-1 epitope has a unique structure comprising the sulfated trisaccharide (HSO(3)-3GlcAbeta1-3Galbeta1-4GlcNAc), and two glucuronyltransferases (GlcAT-P and GlcAT-S) are key enzymes for its biosynthesis. However, the different functional roles of these enzymes in its biosynthesis remain unclear. Recently, we reported that a nonsulfated form of this epitope, which is biosynthesized by GlcAT-S but not by GlcAT-P, is expressed on two metalloproteases in mouse kidney. In this study, we found that a novel glycoprotein carrying the nonsulfated HNK-1 epitope in mouse kidney was enriched in the nuclear fraction. The protein was affinity-purified and identified as laminin-1, and we also confirmed the N-linked oligosaccharide structure including nonsulfated HNK-1 epitope derived from laminin-1 by mass spectrometry. Curiously, immunofluorescence staining of kidney sections revealed that laminin-1 appeared not to be colocalized with the nonsulfated HNK-1 epitope. However, proteinase treatment strengthened the signals of both laminin-1 and the nonsulfated HNK-1 epitope, resulting in overlapping of them. These results indicate that the nonsulfated HNK-1 epitope on laminin-1 is usually embedded and masked in the robust basement membrane in tight association with other proteins. To clarify the associated proteins and the functional role of the carbohydrate epitope, we investigated the interaction between laminin-1 and alpha-dystroglycan through their glycans in mouse kidney using the overlay assay technique. We obtained evidence that glucuronic acid as well as sialic acid inhibited this interaction, suggesting that the nonsulfated HNK-1 epitope on laminin-1 may regulate its binding and play a role in maintenance of the proper structure in the kidney basal lamina.
authors.affiliation: Department of Biological Chemistry, Graduate School of Pharmaceutical Sciences, Kyoto University, Kyoto, Japan.
authors.forename: Yasuhiko
authors.initials: Y
authors.lastname: Kizuka
authors.forename: Kyoko
authors.initials: K
authors.lastname: Kobayashi
authors.forename: Shinako
authors.initials: S
authors.lastname: Kakuda
authors.forename: Yukari
authors.initials: Y
authors.lastname: Nakajima
authors.forename: Satsuki
authors.initials: S
authors.lastname: Itoh
authors.forename: Nana
authors.initials: N
authors.lastname: Kawasaki
authors.forename: Shogo
authors.initials: S
authors.lastname: Oka
mesh_terms: D000818:Animals; D001485:Basement Membrane / Q000737:chemistry / Q000378:metabolism; D018959:CD57 Antigens / Q000378:metabolism* / Q000502:physiology; D002240:Carbohydrate Sequence; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D049030:Dystroglycans / Q000378:metabolism; D000939:Epitopes / Q000378:metabolism / Q000502:physiology; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism / Q000502:physiology; D007668:Kidney / Q000378:metabolism*; D007797:Laminin / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology*; D051379:Mice; D008810:Mice, Inbred C57BL; D008954:Models, Biological; D008969:Molecular Sequence Data; D011485:Protein Binding; D013431:Sulfates / Q000378:metabolism
chemical_list: D018959:CD57 Antigens; D002352:Carrier Proteins; D000939:Epitopes; D006023:Glycoproteins; D007797:Laminin; D013431:Sulfates; C121346:laminin 1; D049030:Dystroglycans