title: Intracellular glutathione pools are heterogeneously concentrated.
abstract: Glutathione is present in millimolar concentrations in the cell, but its relative distribution among cellular compartments remains elusive. We have chosen the endoplasmic reticulum (ER) as an example organelle to study compartment-specific glutathione levels. Using a glutaredoxin sensor (sCGrx1pER), which rapidly and specifically equilibrates with the reduced glutathione (GSH)-glutathione disulfide (GSSG) redox couple with known equilibrium constant, we showed that the [GSH]:[GSSG] ratio in the ER of intact HeLa cells is less than 7:1. Taking into consideration the previously determined value for [GSH](2):[GSSG] in the ER of 83 mM, this translates into a total glutathione concentration in the ER ([GStot]=[GSH]+2[GSSG]) of greater than 15 mM. Since the integrated, intracellular [GStot] was measured as ~7 mM, we conclude the existence of a [GStot] gradient across the ER membrane. A possible homeostatic mechanism by which cytosol-derived glutathione is trapped in the ER is discussed. We propose a high [GStot] as a distinguishing feature of the ER environment compared to the extracellular space.
authors.affiliation: Division of Molecular & Systems Toxicology, Department of Pharmaceutical Sciences, University of Basel, Klingelbergstrasse 50, 4056 Basel, Switzerland.
authors.forename: Davide
authors.initials: D
authors.lastname: Montero
authors.forename: Christine
authors.initials: C
authors.lastname: Tachibana
authors.forename: Jakob
authors.initials: J
authors.lastname: Rahr Winther
authors.forename: Christian
authors.initials: C
authors.lastname: Appenzeller-Herzog
mesh_terms: D003600:Cytosol / Q000378:metabolism; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D054477:Glutaredoxins / Q000235:genetics / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism*; D006367:HeLa Cells; D006706:Homeostasis; D006801:Humans
chemical_list: D054477:Glutaredoxins; D005978:Glutathione
keywords: DTT, Dithiothreitol; EGSH, Half cell reduction potential of glutathione; ER, Endoplasmic reticulum; Endoplasmic reticulum; GSH, Reduced glutathione; GSSG, Glutathione disulfide; Glutaredoxin; Glutathione; NEM, N-ethylmaleimide; OxD, Percentage of oxidation; PDI, Protein disulfide isomerase; PERK, Double stranded RNA-activated protein kinase (PKR)-like ER kinase; RGS, [GSH]:[GSSG]; Redox Homeostasis; Redox compartmentalization; Redox, Reduction–oxidation; Reduction potential; TMM(PEG)12, Maleimide-activated polyethylene glycol; UPR, Unfolded protein response; XBP1, X-box binding protein 1; [GStot], Total glutathione concentration; sCGrx1p, C30S mutant of yeast glutaredoxin 1