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Golgi self-correction generates bioequivalent glycans to preserve cellular homeostasis

View Article: PubMed Central - PubMed

ABSTRACT

Essential biological systems employ self-correcting mechanisms to maintain cellular homeostasis. Mammalian cell function is dynamically regulated by the interaction of cell surface galectins with branched N-glycans. Here we report that N-glycan branching deficiency triggers the Golgi to generate bioequivalent N-glycans that preserve galectin-glycoprotein interactions and cellular homeostasis. Galectins bind N-acetyllactosamine (LacNAc) units within N-glycans initiated from UDP-GlcNAc by the medial-Golgi branching enzymes as well as the trans-Golgi poly-LacNAc extension enzyme β1,3-N-acetylglucosaminyltransferase (B3GNT). Marginally reducing LacNAc content by limiting N-glycans to three branches results in T-cell hyperactivity and autoimmunity; yet further restricting branching does not produce a more hyperactive state. Rather, new poly-LacNAc extension by B3GNT maintains galectin binding and immune homeostasis. Poly-LacNAc extension is triggered by redistribution of unused UDP-GlcNAc from the medial to trans-Golgi via inter-cisternal tubules. These data demonstrate the functional equivalency of structurally dissimilar N-glycans and suggest a self-correcting feature of the Golgi that sustains cellular homeostasis.

Doi:: http://dx.doi.org/10.7554/eLife.14814.001

No MeSH data available.


Related in: MedlinePlus

The hexosamine and N-glycan biosynthetic pathways in mammals.(A) Utilizing the hexosamine pathway product, UDP-GlcNAc, the Golgi enzymes MGAT1, 2, 4a/b, and 5 are responsible for generating mono-, bi-, tri-, and tetra-antennary branched N-glycans, respectively. GlcNAc branches are further modified to create N-acetyllactosamine units, which serve as binding sites for galectins and L-PHA. The alternating actions of galactosyl transferase and B3GNT enzymes produce additional LacNAc units by extending existing branches. MGAT1, 2, 4 and 5 = N-acetylglucosaminyltransferases I, II IV and V; MII/MIIx = mannosidase II/IIx, MI = mannosidase I, SW = swainsonine. Additional structural diversity via addition of sialic acid, fucose, N-acetylgalactosamine and/or sulfate is not shown. (B) T cells isolated from mice of the indicated genotypes were analyzed for L-PHA binding by flow cytometry, gating on CD4+ cells.DOI:http://dx.doi.org/10.7554/eLife.14814.004
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fig1s1: The hexosamine and N-glycan biosynthetic pathways in mammals.(A) Utilizing the hexosamine pathway product, UDP-GlcNAc, the Golgi enzymes MGAT1, 2, 4a/b, and 5 are responsible for generating mono-, bi-, tri-, and tetra-antennary branched N-glycans, respectively. GlcNAc branches are further modified to create N-acetyllactosamine units, which serve as binding sites for galectins and L-PHA. The alternating actions of galactosyl transferase and B3GNT enzymes produce additional LacNAc units by extending existing branches. MGAT1, 2, 4 and 5 = N-acetylglucosaminyltransferases I, II IV and V; MII/MIIx = mannosidase II/IIx, MI = mannosidase I, SW = swainsonine. Additional structural diversity via addition of sialic acid, fucose, N-acetylgalactosamine and/or sulfate is not shown. (B) T cells isolated from mice of the indicated genotypes were analyzed for L-PHA binding by flow cytometry, gating on CD4+ cells.DOI:http://dx.doi.org/10.7554/eLife.14814.004

Mentions: The lattice forms due to the multivalent interactions between extracellular galectins, a family of sugar binding proteins, and the disaccharide N-acetyllactosamine (LacNAc) present on Asn (N)-linked glycans attached to cell surface glycoproteins (Hirabayashi et al., 2002; Brewer et al., 2002; Ahmad et al., 2004). The vast majority of secreted and cell surface proteins are co- or post-translationally modified by the addition of sugars in the ER. As these proteins transit through the ER and Golgi, their glycans undergo dramatic remodeling, generating a vast and heterogeneous array of glycoforms (Kornfeld and Kornfeld, 1985; Schachter, 1991). In the medial Golgi a group of enzymes, MGAT1, 2, 4, and 5, act to produce N-glycans with one, two, three, or four N-acetylglucosamine (GlcNAc) branches (Schachter, 1986). The subsequent addition of galactose by a family of galactosyl transferase enzymes produces the galectin substrate LacNAc (Figure 1—figure supplement 1A). The number of branches depends on the relative activity of the medial Golgi branching enzymes MGAT1, 2, 4, and 5 and the availability of their shared donor substrate UDP-GlcNAc (Lau et al., 2007; Dennis et al., 2009; Grigorian et al., 2007; 2011). Alternating action of β1,3-N-acetylglucosaminyltransferase (B3GNT) and galactosyl transferase enzymes can generate a linear polymer of LacNAc (poly-LacNAc) at any given branch. Although the affinity of galectin binding to a LacNAc monomer is relatively weak, increased LacNAc valency through branching and poly-LacNAc extension can dramatically increase galectin avidity leading to a major impact on cell surface dynamics (Hirabayashi et al., 2002). In T cells for example, galectin - T cell receptor (TCR) interactions directly oppose ligand induced TCR clustering and signaling, thereby negatively regulating T cell development, antigen-dependent T cell growth, and autoimmunity risk.


Golgi self-correction generates bioequivalent glycans to preserve cellular homeostasis
The hexosamine and N-glycan biosynthetic pathways in mammals.(A) Utilizing the hexosamine pathway product, UDP-GlcNAc, the Golgi enzymes MGAT1, 2, 4a/b, and 5 are responsible for generating mono-, bi-, tri-, and tetra-antennary branched N-glycans, respectively. GlcNAc branches are further modified to create N-acetyllactosamine units, which serve as binding sites for galectins and L-PHA. The alternating actions of galactosyl transferase and B3GNT enzymes produce additional LacNAc units by extending existing branches. MGAT1, 2, 4 and 5 = N-acetylglucosaminyltransferases I, II IV and V; MII/MIIx = mannosidase II/IIx, MI = mannosidase I, SW = swainsonine. Additional structural diversity via addition of sialic acid, fucose, N-acetylgalactosamine and/or sulfate is not shown. (B) T cells isolated from mice of the indicated genotypes were analyzed for L-PHA binding by flow cytometry, gating on CD4+ cells.DOI:http://dx.doi.org/10.7554/eLife.14814.004
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4940165&req=5

fig1s1: The hexosamine and N-glycan biosynthetic pathways in mammals.(A) Utilizing the hexosamine pathway product, UDP-GlcNAc, the Golgi enzymes MGAT1, 2, 4a/b, and 5 are responsible for generating mono-, bi-, tri-, and tetra-antennary branched N-glycans, respectively. GlcNAc branches are further modified to create N-acetyllactosamine units, which serve as binding sites for galectins and L-PHA. The alternating actions of galactosyl transferase and B3GNT enzymes produce additional LacNAc units by extending existing branches. MGAT1, 2, 4 and 5 = N-acetylglucosaminyltransferases I, II IV and V; MII/MIIx = mannosidase II/IIx, MI = mannosidase I, SW = swainsonine. Additional structural diversity via addition of sialic acid, fucose, N-acetylgalactosamine and/or sulfate is not shown. (B) T cells isolated from mice of the indicated genotypes were analyzed for L-PHA binding by flow cytometry, gating on CD4+ cells.DOI:http://dx.doi.org/10.7554/eLife.14814.004
Mentions: The lattice forms due to the multivalent interactions between extracellular galectins, a family of sugar binding proteins, and the disaccharide N-acetyllactosamine (LacNAc) present on Asn (N)-linked glycans attached to cell surface glycoproteins (Hirabayashi et al., 2002; Brewer et al., 2002; Ahmad et al., 2004). The vast majority of secreted and cell surface proteins are co- or post-translationally modified by the addition of sugars in the ER. As these proteins transit through the ER and Golgi, their glycans undergo dramatic remodeling, generating a vast and heterogeneous array of glycoforms (Kornfeld and Kornfeld, 1985; Schachter, 1991). In the medial Golgi a group of enzymes, MGAT1, 2, 4, and 5, act to produce N-glycans with one, two, three, or four N-acetylglucosamine (GlcNAc) branches (Schachter, 1986). The subsequent addition of galactose by a family of galactosyl transferase enzymes produces the galectin substrate LacNAc (Figure 1—figure supplement 1A). The number of branches depends on the relative activity of the medial Golgi branching enzymes MGAT1, 2, 4, and 5 and the availability of their shared donor substrate UDP-GlcNAc (Lau et al., 2007; Dennis et al., 2009; Grigorian et al., 2007; 2011). Alternating action of β1,3-N-acetylglucosaminyltransferase (B3GNT) and galactosyl transferase enzymes can generate a linear polymer of LacNAc (poly-LacNAc) at any given branch. Although the affinity of galectin binding to a LacNAc monomer is relatively weak, increased LacNAc valency through branching and poly-LacNAc extension can dramatically increase galectin avidity leading to a major impact on cell surface dynamics (Hirabayashi et al., 2002). In T cells for example, galectin - T cell receptor (TCR) interactions directly oppose ligand induced TCR clustering and signaling, thereby negatively regulating T cell development, antigen-dependent T cell growth, and autoimmunity risk.

View Article: PubMed Central - PubMed

ABSTRACT

Essential biological systems employ self-correcting mechanisms to maintain cellular homeostasis. Mammalian cell function is dynamically regulated by the interaction of cell surface galectins with branched N-glycans. Here we report that N-glycan branching deficiency triggers the Golgi to generate bioequivalent N-glycans that preserve galectin-glycoprotein interactions and cellular homeostasis. Galectins bind N-acetyllactosamine (LacNAc) units within N-glycans initiated from UDP-GlcNAc by the medial-Golgi branching enzymes as well as the trans-Golgi poly-LacNAc extension enzyme β1,3-N-acetylglucosaminyltransferase (B3GNT). Marginally reducing LacNAc content by limiting N-glycans to three branches results in T-cell hyperactivity and autoimmunity; yet further restricting branching does not produce a more hyperactive state. Rather, new poly-LacNAc extension by B3GNT maintains galectin binding and immune homeostasis. Poly-LacNAc extension is triggered by redistribution of unused UDP-GlcNAc from the medial to trans-Golgi via inter-cisternal tubules. These data demonstrate the functional equivalency of structurally dissimilar N-glycans and suggest a self-correcting feature of the Golgi that sustains cellular homeostasis.

Doi:: http://dx.doi.org/10.7554/eLife.14814.001

No MeSH data available.


Related in: MedlinePlus