title: Glucosamine induces ER stress by disrupting lipid-linked oligosaccharide biosynthesis and N-linked protein glycosylation.
abstract: Glucosamine is an essential substrate for N-linked protein glycosylation. However, elevated levels of glucosamine can induce endoplasmic reticulum (ER) stress. Glucosamine-induced ER stress has been implicated in the development of diabetic complications, including atherosclerosis and hepatic steatosis. In this study, we investigate the potential relationship between the effects of glucosamine on lipid-linked oligosaccharide (LLO) biosynthesis, N-linked glycosylation, and ER homeostasis. Mouse embryonic fibroblasts (MEFs) were cultured in the presence of 0-5 mM glucosamine for up to 18 h, and LLO biosynthesis was monitored by fluorescence-assisted carbohydrate electrophoresis. ER stress was determined by quantification of unfolded protein response (UPR) gene expression. We found that exposure of MEFs to ≥1 mM glucosamine significantly impaired the biosynthesis of mature (Glc3Man9GlcNAc2) LLOs before the activation of the UPR, which resulted in the accumulation of an LLO intermediate (Man3GlcNAc2). The addition of 4-phenylbutyric acid (4-PBA), a chemical chaperone, was able to alleviate ER stress but did not rescue LLO biosynthesis. Other ER stress-inducing agents, including dithiothreitol and thapsigargin, had no effect on LLO levels. Together, these data suggest that elevated concentrations of glucosamine induce ER stress by interfering with lipid-linked oligosaccharide biosynthesis and N-linked glycosylation. We hypothesize that this pathway represents a causative link between hyperglycemia and the development of diabetic complications.
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, Ontario, Canada.|Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Daniel R
authors.initials: DR
authors.lastname: Beriault
authors.affiliation: Department of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario, Canada.|Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Vi T
authors.initials: VT
authors.lastname: Dang
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Lexy H
authors.initials: LH
authors.lastname: Zhong
authors.affiliation: Department of Chemistry and Chemical Biology, McMaster University, Hamilton, Ontario, Canada.|Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Christina I
authors.initials: CI
authors.lastname: Petlura
authors.affiliation: Department of Medicine McMaster University, Hamilton, Ontario, Canada; and.|Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Cameron S
authors.initials: CS
authors.lastname: McAlpine
authors.affiliation: Thrombosis and Atherosclerosis Research Institute, McMaster University, Hamilton, Ontario, Canada.
authors.forename: Yuanyuan
authors.initials: Y
authors.lastname: Shi
authors.forename: Geoff H
authors.initials: GH
authors.lastname: Werstuck
mesh_terms: D000818:Animals; D002460:Cell Line; D004229:Dithiothreitol / Q000494:pharmacology; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects* / Q000378:metabolism; D005944:Glucosamine / Q000494:pharmacology*; D006031:Glycosylation / Q000187:drug effects*; D008070:Lipopolysaccharides / Q000096:biosynthesis*; D051379:Mice; D010654:Phenylbutyrates / Q000494:pharmacology; D019284:Thapsigargin / Q000494:pharmacology; D056811:Unfolded Protein Response / Q000187:drug effects
chemical_list: D004791:Enzyme Inhibitors; D008070:Lipopolysaccharides; D010654:Phenylbutyrates; C023023:lipid-linked oligosaccharides; D019284:Thapsigargin; C075773:4-phenylbutyric acid; D005944:Glucosamine; D004229:Dithiothreitol
keywords: endoplasmic reticulum