title: Disulphide production by Ero1α-PDI relay is rapid and effectively regulated.
abstract: The molecular networks that control endoplasmic reticulum (ER) redox conditions in mammalian cells are incompletely understood. Here, we show that after reductive challenge the ER steady-state disulphide content is restored on a time scale of seconds. Both the oxidase Ero1α and the oxidoreductase protein disulphide isomerase (PDI) strongly contribute to the rapid recovery kinetics, but experiments in ERO1-deficient cells indicate the existence of parallel pathways for disulphide generation. We find PDI to be the main substrate of Ero1α, and mixed-disulphide complexes of Ero1 primarily form with PDI, to a lesser extent with the PDI-family members ERp57 and ERp72, but are not detectable with another homologue TMX3. We also show for the first time that the oxidation level of PDIs and glutathione is precisely regulated. Apparently, this is achieved neither through ER import of thiols nor by transport of disulphides to the Golgi apparatus. Instead, our data suggest that a dynamic equilibrium between Ero1- and glutathione disulphide-mediated oxidation of PDIs constitutes an important element of ER redox homeostasis.
authors.affiliation: Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Christian
authors.initials: C
authors.lastname: Appenzeller-Herzog
authors.forename: Jan
authors.initials: J
authors.lastname: Riemer
authors.forename: Ester
authors.initials: E
authors.lastname: Zito
authors.forename: King-Tung
authors.initials: KT
authors.lastname: Chin
authors.forename: David
authors.initials: D
authors.lastname: Ron
authors.forename: Martin
authors.initials: M
authors.lastname: Spiess
authors.forename: Lars
authors.initials: L
authors.lastname: Ellgaard
mesh_terms: D002478:Cells, Cultured; D003208:Concanavalin A / Q000302:isolation & purification; D017931:DNA Primers / Q000235:genetics; D003720:Densitometry; D004220:Disulfides / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D047468:Immunoprecipitation; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000378:metabolism*; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D014162:Transfection
chemical_list: D017931:DNA Primers; D004220:Disulfides; D008562:Membrane Glycoproteins; D003208:Concanavalin A; C405327:ERO1A protein, human; D010088:Oxidoreductases; D019704:Protein Disulfide-Isomerases; C498879:TMX3 protein, human; D005978:Glutathione
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