Dataset Information

Glycosylation analysis of TNF-alpha-induced insulin resistance

ABSTRACT: Glycoproteomics, TNF-alpha, membrane proteomics, insulin resistance, adipocyte, SILAC Insulin resistance (IR) is a complex process arising from both environmental and genetic perturbations and leads to a variety of diseases including type-2 diabetes (T2D). The expansion of adipose tissue during obesity results in secretion of proinflammatory cytokines which significantly impairs insulin sensitivity throughout the entire body. Inflammation modulates glycosylation in a variety of cell types however, an investigation of changes in glycosylation following inflammation-induced IR in adipocytes has not been performed. In the present study, we performed a quantitative N-glycomic analysis of TNF-alpha induced IR in adipocytes and identified the regulation of specific N-glycans including an increase in terminal di-galactose- and decrease in di-sialic acid-containing glycans with alpha-2,3 linkages. Quantitative analysis of the membrane-associated proteome in TNF-alpha treated adipocytes identified the regulation of specific glycosyltransferases and glycosidases which correlated with the regulated N-glycans. This included the up- and down-regulation of B4GalT5 and ST3Gal6, respectively at both the protein and mRNA level. To identify the protein and glycosylation sites modified with these regulated N-glycans, we performed a site-specific quantitative glycoproteomic analysis of enriched N-glycopeptides from TNF-alpha treated adipocytes with and without deglycosylation. The combined workflow provided a relative quantification of changes in protein abundance verses N-glycosylation occupancy verses site-specific N-glycans on a proteome-wide level. This revealed the modulation of a subset of N-glycan compositions on specific proteins including those previously associated with insulin-stimulated GLUT4 trafficking to the plasma membrane. Finally, knockdown of B4GalT5 with siRNA and analysis of released N-glycans resulted in the down-regulation of di-galactose containing glycans, confirming the involvement of this enzyme in the TNF-alpha regulated N-glycome.

INSTRUMENT(S): Orbitrap Fusion ETD, Q Exactive

ORGANISM(S): Mus Musculus (mouse)

TISSUE(S): Fat Cell

SUBMITTER: Benjamin Parker

LAB HEAD: Benjamin Parker

PROVIDER: PXD002435 | Pride | 2015-11-09

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	150219_QEplus_BP_NGlyco_rep1_FT_1.raw	Raw
	150219_QEplus_BP_NGlyco_rep1_FT_10.raw	Raw
	150219_QEplus_BP_NGlyco_rep1_FT_11.raw	Raw
	150219_QEplus_BP_NGlyco_rep1_FT_12.raw	Raw
	150219_QEplus_BP_NGlyco_rep1_FT_2.raw	Raw

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Publications

Terminal Galactosylation and Sialylation Switching on Membrane Glycoproteins upon TNF-Alpha-Induced Insulin Resistance in Adipocytes.

Parker Benjamin L BL Thaysen-Andersen Morten M Fazakerley Daniel J DJ Holliday Mira M Packer Nicolle H NH James David E DE

Molecular & cellular proteomics : MCP 20151104 1

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 ...[more]

PMID: 26537798

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