Dataset Information

Developmental expression of the neuron-specific N-acetylglucosaminyltransferase Vb (GnT-Vb/IX) and identification of its in vivo glycan products in comparison with those of its paralog, GnT-V.

ABSTRACT: The severe phenotypic effects of altered glycosylation in the congenital muscular dystrophies, including Walker-Warburg syndrome, muscle-eye-brain disease, Fukuyama congenital muscular dystrophy, and congenital muscular dystrophy 1D, are caused by mutations resulting in altered glycans linked to proteins through O-linked mannose. A glycosyltransferase that branches O-Man, N-acetylglucosaminyltransferase Vb (GnT-Vb), is highly expressed in neural tissues. To understand the expression and function of GnT-Vb, we studied its expression during neuromorphogenesis and generated GnT-Vb null mice. A paralog of GnT-Vb, N-acetylglucosaminyltransferase (GnT-V), is expressed in many tissues and brain, synthesizing N-linked, ?1,6-branched glycans, but its ability to synthesize O-mannosyl-branched glycans is unknown; conversely, although GnT-Vb can synthesize N-linked glycans in vitro, its contribution to their synthesis in vivo is unknown. Our results showed that deleting both GnT-V and GnT-Vb results in the total loss of both N-linked and O-Man-linked ?1,6-branched glycans. GnT-V null brains lacked N-linked, ?1,6-glycans but had normal levels of O-Man ?1,6-branched structures, showing that GnT-Vb could not compensate for the loss of GnT-V. By contrast, GnT-Vb null brains contained normal levels of N-linked ?1,6-glycans but low levels of some O-Man ?1,6-branched glycans. Therefore, GnT-V could partially compensate for GnT-Vb activity in vivo. We found no apparent change in ?-dystroglycan binding of glycan-specific antibody IIH6C4 or binding to laminin in GnT-Vb null mice. These results demonstrate that GnT-V is involved in synthesizing branched O-mannosyl glycans in brain, but the function of these branched O-mannosyl structures is unresolved using mice that lack these glycosyltransferases.

SUBMITTER: Lee JK

PROVIDER: S-EPMC3436567 | biostudies-literature | 2012 Aug

REPOSITORIES: biostudies-literature

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Developmental expression of the neuron-specific N-acetylglucosaminyltransferase Vb (GnT-Vb/IX) and identification of its in vivo glycan products in comparison with those of its paralog, GnT-V.

Lee Jin Kyu JK Matthews Russell T RT Lim Jae-Min JM Swanier Kiara K Wells Lance L Pierce J Michael JM

The Journal of biological chemistry 20120619 34

The severe phenotypic effects of altered glycosylation in the congenital muscular dystrophies, including Walker-Warburg syndrome, muscle-eye-brain disease, Fukuyama congenital muscular dystrophy, and congenital muscular dystrophy 1D, are caused by mutations resulting in altered glycans linked to proteins through O-linked mannose. A glycosyltransferase that branches O-Man, N-acetylglucosaminyltransferase Vb (GnT-Vb), is highly expressed in neural tissues. To understand the expression and function ...[more]

PMID: 22715095

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Project description:N-Acetylglucosaminlyltransferase-V (GnT-V) synthesizes GlcNAcbeta1,6Man branched N-glycans both in vitro and in vivo. A paralog, GnT-Vb (or GnT-IX), has also been shown to synthesize both GlcNAcbeta1,6Man branched N- and O-glycans. GnT-V is expressed in most human and rodent tissues while GnT-Vb expression is limited mainly to neural tissue and testes. It is of interest, therefore, to compare the catalytic properties and reaction kinetics of these sister enzymes. The results demonstrate that while GnT-V was fully active without exogenous cation and in the presence of EDTA, the activity of GnT-Vb was stimulated over 4-fold in the presence of 10 mM Mn(++). The pH optimum for GnT-V was in the range of 6.5-7.0, while that of GnT-Vb was 8.0. common for glycosyltransferases active in brain. Both enzymes transferred GlcNAcbeta1,6 to the Man residue of the GlcNAcbeta1,2Man moiety of glycan substrates, and both enzymes acted effectively on a synthetic GlcNAcbeta1,2Manalpha1,2Glc-O-octyl trisaccharide acceptor. Moreover, although both enzymes utilized an N-linked asialo-agalacto-biantennary glycan as an acceptor, GnT-Vb displayed an almost 2.5-fold higher apparent K(m) value compared to GnT-V. Conversely, GnT-Vb very efficiently glycosylated a synthetic glycopeptide, Ac-H(2)N-Val-Glu-Pro-(GlcNAcbeta1,2-Man-O-)Thr-Ala-Val-CO-Ac, while GnT-V showed relatively poor activity toward this O-Man-linked glycopeptide acceptor, with a K(m) value of 20-fold higher than that of GnT-Vb. When the N-linked asialo-agalacto-biantennary glycan acceptor was utilized with GnT-Vb, the expected triantennary beta1,6-branched product was observed up to 8 h incubation. An additional product with two beta1,6-linked GlcNAc resides, however, was observed after prolonged (>8 h) incubation, consistent with an earlier report. This unusual tetraantennary product was observed with GnT-Vb only after substantial accumulation of the first triantennary product and not during the early stages of incubation.

Dataset Information

Developmental expression of the neuron-specific N-acetylglucosaminyltransferase Vb (GnT-Vb/IX) and identification of its in vivo glycan products in comparison with those of its paralog, GnT-V.

Publications

Developmental expression of the neuron-specific N-acetylglucosaminyltransferase Vb (GnT-Vb/IX) and identification of its in vivo glycan products in comparison with those of its paralog, GnT-V.

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