Molecular Cloning and Expression of a Second Glucuronyltransferase Involved in the Biosynthesis of the HNK-1 Carbohydrate Epitope

Seiki, Takashi; Oka, Shogo; Terayama, Koji; Imiya, Kimiyuki; Kawasaki, Toshisuke

doi:10.1006/bbrc.1999.0151

Cited by 70 publications

(36 citation statements)

References 34 publications

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“…GlcAT-S shows 89% homology with the rat protein, with the highest identity being found in the C-terminal catalytic domain, which contains four highly conserved regions, named motifs I-IV, as previously reported elsewhere (see Fig. 2) (Seiki et al 1999). The high sequence conservation between the rat and the human GlcAT-S strongly suggests conserved functions of this gene in both species.…”

Section: Resultssupporting

confidence: 82%

Cloning, characterization, and chromosome mapping of the human GlcAT-S gene

Marcos¹,

Borrego²,

Antiñolo³

2002

J Hum Genet

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Section: Resultssupporting

confidence: 82%

Cloning, characterization, and chromosome mapping of the human GlcAT-S gene

Marcos¹,

Borrego²,

Antiñolo³

2002

J Hum Genet

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“…Two glucuronyltransferases (GlcAT-P and GlcAT-S) have so far been identified as HNK-1 biosynthetic enzymes (7)(8)(9)(10)(11). We have generated GlcAT-P gene-deficient mice and demonstrated that almost all of the HNK-1 carbohydrate in the brain is synthesized by GlcAT-P because of the absence of the HNK-1 carbohydrate in GlcAT-P gene-deficient mouse brain (12).…”

Section: Discussionmentioning

confidence: 99%

“…Therefore, glucuronyltransferase(s) and sulfotransferase(s) are supposed to be key enzymes for the biosynthesis of this carbohydrate (6). Recently, we cloned two glucuronyltransferases (GlcAT-P and GlcAT-S) that are involved in the biosynthesis of the HNK-1 carbohydrate from rat, mouse, and human (7)(8)(9)(10)(11). To elucidate the function of the HNK-1 carbohydrate, we generated mice with a targeted deletion of the GlcAT-P gene (12).…”

mentioning

confidence: 99%

A Non-sulfated Form of the HNK-1 Carbohydrate Is Expressed in Mouse Kidney

Tagawa

Kizuka

Ikeda³

et al. 2005

Journal of Biological Chemistry

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The HNK-1 carbohydrate, which is recognized by anti-HNK-1 antibody, is well known to be expressed predominantly in the nervous system. The characteristic structural feature of the HNK-1 carbohydrate is 3-sulfoglucuronyl residues attached to lactosamine structures (Gal␤1-4GlcNAc) on glycoproteins and glycolipids. The biosynthesis of the HNK-1 carbohydrate is regulated mainly by two glucuronyltransferases (GlcAT-P and GlcAT-S) and a sulfotransferase. In this study, we found that GlcAT-S mRNA was expressed at higher levels in the kidney than in the brain, but that both GlcAT-P and HNK-1 sulfotransferase mRNAs, which were expressed at high levels in the brain, were not detected in the kidney. These results suggested that the HNK-1 carbohydrate without sulfate (non-sulfated HNK-1 carbohydrate) is expressed in the kidney. We substantiated this hypothesis using two different monoclonal antibodies: one (anti-HNK-1 antibody) requires sulfate on glucuronyl residues for its binding, and the other (antibody M6749) does not. Western blot analyses of mouse kidney revealed that two major bands (80 and 140 kDa) were detected with antibody M6749, but not with anti-HNK-1 antibody. The 80-and 140-kDa band materials were identified as meprin ␣ and CD13/aminopeptidase N, respectively. We also confirmed the presence of the non-sulfated HNK-1 carbohydrate on N-linked oligosaccharides by multistage tandem mass spectrometry. Immunofluorescence staining with antibody M6749 revealed that the non-sulfated HNK-1 carbohydrate was expressed predominantly on the apical membranes of the proximal tubules in the cortex and was also detected in the thin ascending limb in the inner medulla. This is the first study indicating the presence of the non-sulfated HNK-1 carbohydrate being synthesized by GlcAT-S in the kidney. The results presented here constitute novel knowledge concerning the function of the HNK-1 carbohydrate.Glycosylation is one of the major post-translational protein modifications that play important roles in a variety of cellular functions, including recognition and adhesion. We have been interested in the HNK-1 (human natural killer-1) carbohydrate, which is recognized by anti-HNK-1 monoclonal antibody (1). The carbohydrate is expressed predominantly in the nervous system in a wide range of species, and its expression is spatially and temporally regulated during development of the nervous system (2, 3). The characteristic structural feature of the carbohydrate is a sulfated trisaccharide, HSO 3 -3GlcA␤1-3Gal␤1-4GlcNAc 1 (4, 5), and the inner structure, Gal␤1-4GlcNAc, is commonly found on various glycoproteins and glycolipids. Therefore, glucuronyltransferase(s) and sulfotransferase(s) are supposed to be key enzymes for the biosynthesis of this carbohydrate (6). Recently, we cloned two glucuronyltransferases (GlcAT-P and GlcAT-S) that are involved in the biosynthesis of the HNK-1 carbohydrate from rat, mouse, and human (7-11). To elucidate the function of the HNK-1 carbohydrate, we generated mice with a targeted deletion of the GlcAT...

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“…HNK-1 carbohydrate has a unique structure, including a sulfated trisaccharide (HSO 3 -3GlcA␤1-3Gal␤1-4GlcNAc-) (9,10), and is biosynthesized by the successive actions of ␤-1,4-galactosyltransferase (␤4GalT), a glucuronyltransferase (GlcAT-P or GlcAT-S), and a sulfotransferase (HNK-1ST) (11)(12)(13). We previously generated mice lacking the gene for GlcAT-P, a major glucuronyltransferase in the nervous system (14).…”

mentioning

confidence: 99%

Specific Enzyme Complex of β-1,4-Galactosyltransferase-II and Glucuronyltransferase-P Facilitates Biosynthesis of N-linked Human Natural Killer-1 (HNK-1) Carbohydrate

Kouno

Kizuka

Nakagawa

et al. 2011

Journal of Biological Chemistry

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Human natural killer-1 (HNK-1) carbohydrate is highly expressed in the nervous system and is involved in synaptic plasticity and dendritic spine maturation. This unique carbohydrate, consisting of a sulfated trisaccharide (HSO 3 -3GlcA␤1-3Gal␤1-4GlcNAc-), is biosynthesized by the successive actions of ␤-1,4-galactosyltransferase (␤4GalT), glucuronyltransferase (GlcAT-P and GlcAT-S), and sulfotransferase (HNK-1ST). A previous study showed that mice lacking ␤4GalT-II, one of seven ␤4GalTs, exhibited a dramatic loss of HNK-1 expression in the brain, although ␤4GalT-I-deficient mice did not. Here, we investigated the underlying molecular mechanism of the regulation of HNK-1 expression. First, focusing on a major HNK-1 carrier, neural cell adhesion molecule, we found that reduced expression of an N-linked HNK-1 carbohydrate caused by a deficiency of ␤4GalT-II is not likely due to a general loss of the ␤1,4-galactose residue as an acceptor for GlcAT-P. Instead, we demonstrated by co-immunoprecipitation and endoplasmic reticulum-retention analyses using Neuro2a (N2a) cells that ␤4GalT-II physically and specifically associates with GlcAT-P. In addition, we revealed by pulldown assay that Golgi luminal domains of ␤4GalT-II and GlcAT-P are sufficient for the complex to form. With an in vitro assay system, we produced the evidence that the kinetic efficiency k cat /K m of GlcAT-P in the presence of ␤4GalT-II was increased about 2.5-fold compared with that in the absence of ␤4GalT-II. Finally, we showed that co-expression of ␤4GalT-II and GlcAT-P increased HNK-1 expression on various glycoproteins in N2a cells, including neural cell adhesion molecule. These results indicate that the specific enzyme complex of ␤4GalT-II with GlcAT-P plays an important role in the biosynthesis of HNK-1 carbohydrate.

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Molecular Cloning and Expression of a Second Glucuronyltransferase Involved in the Biosynthesis of the HNK-1 Carbohydrate Epitope

Cited by 70 publications

References 34 publications

Cloning, characterization, and chromosome mapping of the human GlcAT-S gene

Cloning, characterization, and chromosome mapping of the human GlcAT-S gene

A Non-sulfated Form of the HNK-1 Carbohydrate Is Expressed in Mouse Kidney

Specific Enzyme Complex of β-1,4-Galactosyltransferase-II and Glucuronyltransferase-P Facilitates Biosynthesis of N-linked Human Natural Killer-1 (HNK-1) Carbohydrate

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