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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

TCR signaling and cellular UDP-GlcNAc levels modulate the degree of poly-LacNAc induced by branching Deficiency.(A–C) Jurkat T cells were treated with SW for the indicated times and analyzed for lectin binding by flow cytometry. (D and E) Jurkat T cells were treated as indicated for 72 hr and analyzed for LEA binding by flow cytometry. (F and G) Resting primary human T cells were treated as indicated and analyzed for LEA (F) or L-PHA (G) binding by flow cytometry. NS, not significant; *p<0.05; **p<0.01; ***p<0.001 (unpaired two-tailed t-test with Welch’s (D–G) and Bonferroni correction (D–G)). Data show one experiment representative of at least three independent experiments. Error bars indicate mean ± s.e.m.DOI:http://dx.doi.org/10.7554/eLife.14814.012
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fig4s1: TCR signaling and cellular UDP-GlcNAc levels modulate the degree of poly-LacNAc induced by branching Deficiency.(A–C) Jurkat T cells were treated with SW for the indicated times and analyzed for lectin binding by flow cytometry. (D and E) Jurkat T cells were treated as indicated for 72 hr and analyzed for LEA binding by flow cytometry. (F and G) Resting primary human T cells were treated as indicated and analyzed for LEA (F) or L-PHA (G) binding by flow cytometry. NS, not significant; *p<0.05; **p<0.01; ***p<0.001 (unpaired two-tailed t-test with Welch’s (D–G) and Bonferroni correction (D–G)). Data show one experiment representative of at least three independent experiments. Error bars indicate mean ± s.e.m.DOI:http://dx.doi.org/10.7554/eLife.14814.012

Mentions: As the galectin-glycoprotein lattice negatively regulates TCR signaling and TCR signaling promotes lattice strength (Demetriou et al., 2001; Chen et al., 2009), we hypothesized that LacNAc homeostasis may result from a feedback loop linking TCR signaling and cell surface LacNAc content. Such a mechanism, which depends on a cell surface sensor of Golgi activity/branching implies a temporal lag phase during which a defect is detected, a signal is sent, and then the Golgi generates the proper response. With this prediction in mind, Jurkat T cells were treated with SW for various times and analyzed for changes in cell surface glycosylation by L-PHA, LEA, and Concanavalin A (ConA), the latter a plant lectin that binds high-mannose structures increased by SW treatment (Figure 4—figure supplement 1A–C). Although the increase in LEA binding trailed slightly behind an increase in ConA binding, it began to increase within ~1.5 hr of SW treatment, indicating an almost immediate compensatory response. Loss of L-PHA staining exhibited the smallest slope, possibly reflecting the preferential cell surface retention of highly branched glycoproteins (Figure 4—figure supplement 1C).


Golgi self-correction generates bioequivalent glycans to preserve cellular homeostasis
TCR signaling and cellular UDP-GlcNAc levels modulate the degree of poly-LacNAc induced by branching Deficiency.(A–C) Jurkat T cells were treated with SW for the indicated times and analyzed for lectin binding by flow cytometry. (D and E) Jurkat T cells were treated as indicated for 72 hr and analyzed for LEA binding by flow cytometry. (F and G) Resting primary human T cells were treated as indicated and analyzed for LEA (F) or L-PHA (G) binding by flow cytometry. NS, not significant; *p<0.05; **p<0.01; ***p<0.001 (unpaired two-tailed t-test with Welch’s (D–G) and Bonferroni correction (D–G)). Data show one experiment representative of at least three independent experiments. Error bars indicate mean ± s.e.m.DOI:http://dx.doi.org/10.7554/eLife.14814.012
© Copyright Policy
Related In: Results  -  Collection

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fig4s1: TCR signaling and cellular UDP-GlcNAc levels modulate the degree of poly-LacNAc induced by branching Deficiency.(A–C) Jurkat T cells were treated with SW for the indicated times and analyzed for lectin binding by flow cytometry. (D and E) Jurkat T cells were treated as indicated for 72 hr and analyzed for LEA binding by flow cytometry. (F and G) Resting primary human T cells were treated as indicated and analyzed for LEA (F) or L-PHA (G) binding by flow cytometry. NS, not significant; *p<0.05; **p<0.01; ***p<0.001 (unpaired two-tailed t-test with Welch’s (D–G) and Bonferroni correction (D–G)). Data show one experiment representative of at least three independent experiments. Error bars indicate mean ± s.e.m.DOI:http://dx.doi.org/10.7554/eLife.14814.012
Mentions: As the galectin-glycoprotein lattice negatively regulates TCR signaling and TCR signaling promotes lattice strength (Demetriou et al., 2001; Chen et al., 2009), we hypothesized that LacNAc homeostasis may result from a feedback loop linking TCR signaling and cell surface LacNAc content. Such a mechanism, which depends on a cell surface sensor of Golgi activity/branching implies a temporal lag phase during which a defect is detected, a signal is sent, and then the Golgi generates the proper response. With this prediction in mind, Jurkat T cells were treated with SW for various times and analyzed for changes in cell surface glycosylation by L-PHA, LEA, and Concanavalin A (ConA), the latter a plant lectin that binds high-mannose structures increased by SW treatment (Figure 4—figure supplement 1A–C). Although the increase in LEA binding trailed slightly behind an increase in ConA binding, it began to increase within ~1.5 hr of SW treatment, indicating an almost immediate compensatory response. Loss of L-PHA staining exhibited the smallest slope, possibly reflecting the preferential cell surface retention of highly branched glycoproteins (Figure 4—figure supplement 1C).

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 &beta;1,3-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