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Terminal galactosylation and sialylation switching on membrane glycoproteins upon TNF-alpha-induced insulin resistance in adipocytes

Reputable Mentor II
Reputable Mentor II
Benjamin L. Parker, Morten Thaysen-Andersen, Daniel J. Fazakerley, Mira Holliday, Nicolle H. Packer, and David E. James
Mol Cell Proteomics mcp.M115.054221. First Published on November 4, 2015, doi:10.1074/mcp.M115.054221
Insulin resistance (IR) is a complex pathophysiological state that arises from both environmental and genetic perturbations and leads to a variety of diseases including type-2 diabetes (T2D). Obesity is associated with enhanced adipose tissue inflammation, which may play a role in disease progression. Inflammation modulates protein glycosylation in a variety of cell types and this has been associated with biological dysregulation. Here we have examined the effects of an inflammatory insult on protein glycosylation in adipocytes. We performed quantitative N-glycome profiling of membrane proteins derived from mouse 3T3-L1 adipocytes that had been incubated with or without the proinflammatory cytokine TNF-alpha to induce IR. We identified the regulation of specific terminal N-glycan epitopes, including an increase in terminal di-galactose- and a decrease in biantennary alpha-2,3-sialoglycans. The altered N-glycosylation of TNF-alpha treated adipocytes correlated with the regulation of specific glycosyltransferases including the up-regulation of B4GalT5 and Ggta1 galactosyltransferases and down-regulation of ST3Gal6 sialyltransferase. Knockdown of B4GalT5 down-regulated the terminal di-galactose N-glycans, confirming the involvement of this enzyme in the TNF-alpha regulated N-glycome. SILAC-based quantitative glycoproteomics of enriched N-glycopeptides with and without deglycosylation were used to identify the protein and glycosylation sites modified with these regulated N-glycans. The combined proteome and glycoproteome workflow provided a relative quantification of changes in protein abundance versus N-glycosylation occupancy versus site-specific N-glycans on a proteome-wide level. This revealed the modulation of N-glycosylation on specific proteins in IR, including those previously associated with insulin-stimulated GLUT4 trafficking to the plasma membrane.
Charles Perkins Centre, School of Molecular Bioscience, University of Sydney, Sydney, Australia Department of Chemistry and Biomolecular Sciences, Macquarie University, Sydney, Australia School of Medicine, University of Sydney, Sydney, Australia
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‎10-15-2021 11:06 AM
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