title: Endoplasmic reticulum redox state is not perturbed by pharmacological or pathological endoplasmic reticulum stress in live pancreatic β-cells.
abstract: Accumulation of unfolded, misfolded and aggregated proteins in the endoplasmic reticulum (ER) causes ER stress. ER stress can result from physiological situations such as acute increases in secretory protein biosynthesis or pathological conditions that perturb ER homeostasis such as alterations in the ER redox state. Here we monitored ER redox together with transcriptional output of the Unfolded Protein Response (UPR) in INS-1 insulinoma cells stably expressing eroGFP (ER-redox-sensor) and mCherry protein driven by a GRP78 promoter (UPR-sensor). Live cell imaging, flow cytometry and biochemical characterization were used to examine these parameters in response to various conditions known to induce ER stress. As expected, treatment of the cells with the reducing agent dithiothreitol caused a decrease in the oxidation state of the ER accompanied by an increase in XBP-1 splicing. Unexpectedly however, other treatments including tunicamycin, thapsigargin, DL-homocysteine, elevated free fatty acids or high glucose had essentially no influence on the ER redox state, despite inducing ER stress. Comparable results were obtained with dispersed rat islet cells expressing eroGFP. Thus, unlike in yeast cells, ER stress in pancreatic β-cells is not associated with a more reducing ER environment.
authors.affiliation: Division of Cellular and Molecular Biology, Toronto General Research Institute, University Health Network, Toronto, Ontario, Canada.
authors.forename: Irmgard
authors.initials: I
authors.lastname: Schuiki
authors.forename: Liling
authors.initials: L
authors.lastname: Zhang
authors.forename: Allen
authors.initials: A
authors.lastname: Volchuk
mesh_terms: D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D003920:Diabetes Mellitus / Q000378:metabolism / Q000473:pathology; D004229:Dithiothreitol / Q000494:pharmacology*; D004721:Endoplasmic Reticulum / Q000187:drug effects* / Q000378:metabolism*; D059865:Endoplasmic Reticulum Stress / Q000187:drug effects*; D005230:Fatty Acids, Nonesterified / Q000494:pharmacology; D005434:Flow Cytometry; D005947:Glucose / Q000494:pharmacology; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006710:Homocysteine / Q000494:pharmacology; D050417:Insulin-Secreting Cells / Q000187:drug effects / Q000378:metabolism* / Q000473:pathology*; D008164:Luminescent Proteins / Q000378:metabolism; D009765:Obesity / Q000378:metabolism / Q000473:pathology; D010084:Oxidation-Reduction / Q000187:drug effects; D051381:Rats; D000097573:Red Fluorescent Protein
chemical_list: D004229:Dithiothreitol; D005230:Fatty Acids, Nonesterified; D005947:Glucose; D049452:Green Fluorescent Proteins; D006710:Homocysteine; D008164:Luminescent Proteins; D000097573:Red Fluorescent Protein