Significance: The thioredoxin (TRX) and glutathione (GSH) pathways are universally conserved thiol-reductase systems that drive an array of cellular functions involving reversible disulfide formation. Here we consider these pathways in Saccharomyces cerevisiae, focusing on their cell compartment-specific functions, as well as the mechanisms that explain extreme differences of redox states between compartments. Recent Advances: Recent work leads to a model in which the yeast TRX and GSH pathways are not redundant, in contrast to Escherichia coli. The cytosol possesses full sets of both pathways, of which the TRX pathway is dominant, while the GSH pathway acts as back up of the former. The mitochondrial matrix also possesses entire sets of both pathways, in which the GSH pathway has major role in redox control. In both compartments, GSH has also nonredox functions in iron metabolism, essential for viability. The endoplasmic reticulum (ER) and mitochondrial intermembrane space (IMS) are sites of intense thiol oxidation, but except GSH lack thiol-reductase pathways. Critical Issues: What are the thiol-redox links between compartments? Mitochondria are totally independent, and insulated from the other compartments. The cytosol is also totally independent, but also provides reducing power to the ER and IMS, possibly by ways of reduced and oxidized GSH entering and exiting these compartments. Future Directions: Identifying the mechanisms regulating fluxes of GSH and oxidized glutathione between cytosol and ER, IMS, and possibly also peroxisomes, vacuole is needed to establish the proposed model of eukaryotic thiol-redox homeostasis, which should facilitate exploration of this system in mammals and plants. © 2013 Mary Ann Liebert, Inc.

In the endoplasmic reticulum (ER), Ero1 catalyzes disulfide bond formation and promotes glutathione (GSH) oxidation to GSSG. Since GSSG cannot be reduced in the ER, maintenance of the ER glutathione redox state and levels likely depends on ER glutathione import and GSSG export. We used quantitative GSH and GSSG biosensors to monitor glutathione import into the ER of yeast cells. We found that glutathione enters the ER by facilitated diffusion through the Sec61 protein-conducting channel, while oxidized Bip (Kar2) inhibits transport. Increased ER glutathione import triggers H₂O₂-dependent Bip oxidation through Ero1 reductive activation, which inhibits glutathione import in a negative regulatory loop. During ER stress, transport is activated by UPR-dependent Ero1 induction, and cytosolic glutathione levels increase. Thus, the ER redox poise is tuned by reciprocal control of glutathione import and Ero1 activation. The ER protein-conducting channel is permeable to small molecules, provided the driving force of a concentration gradient. Ponsero et al. show that cytosol-to-ER transport of glutathione proceeds via facilitated diffusion through Sec61. Upon import, glutathione activates Ero1 by reduction, causing Bip oxidation and inhibition of glutathione transport. Coupling of glutathione ER import to Ero1 activation provides a basis for glutathione ER redox poise maintenance. © 2017 Elsevier Inc.

Glutathione contributes to thiol-redox control and to extra-mitochondrial irong-sulphur cluster (ISC) maturation. To determine the physiological importance of these functions and sort out those that account for the GSH requirement for viability, we performed a comprehensive analysis of yeast cells depleted of or containing toxic levels of GSH. Both conditions triggered an intense iron starvation-like response and impaired the activity of extra-mitochondrial ISC enzymes but did not impact thiol-redox maintenance, except for high glutathione levels that altered oxidative protein folding in the endoplasmic reticulum. While iron partially rescued the ISC maturation and growth defects of GSH-depleted cells, genetic experiments indicated that unlike thioredoxin, glutathione could not support by itself the thiol-redox duties of the cell. We propose that glutathione is essential by its requirement in ISC assembly, but only serves as a thioredoxin backup in cytosolic thiol-redox maintenance. Glutathione-high physiological levels are thus meant to insulate its cytosolic function in iron metabolism from variations of its concentration during redox stresses, a model challenging the traditional view of it as prime actor in thiol-redox control. © 2011 European Molecular Biology Organization

The oxidation of the cysteine (Cys) residue to sulfenic (–S–OH), disulfide (–S–S–), or S-nitroso (S–NO) forms are thought to be a posttranslational modifications that regulate protein function. However, despite a few solid examples of its occurrence, thiol-redox regulation of protein function is still debated and often seen as an exotic phenomenon. A systematic and exhaustive characterization of all oxidized Cys residues, an experimental approach called redox proteomics or redoxome analysis, should help establish the physiological scope of Cys residue oxidation and give clues to its mechanisms. Redox proteomics still remains a technical challenge, mainly because of the labile nature of thiol-redox reactions and the lack of tools to directly detect the modified residues. Here we consider recent technical advances in redox proteomics, focusing on a gel-based fluorescent method and on the shotgun OxICAT technique.

Yeast lacks dedicated photoreceptors; however, blue light still causes pronounced oscillations of the transcription factor Msn2 into and out of the nucleus. Here we show that this poorly understood phenomenon is initiated by a peroxisomal oxidase, which converts light into a hydrogen peroxide (H 2 O 2) signal that is sensed by the peroxiredoxin Tsa1 and transduced to thioredoxin, to counteract PKA-dependent Msn2 phosphorylation. Upon H 2 O 2, the nuclear retention of PKA catalytic subunits, which contributes to delayed Msn2 nuclear concentration, is antagonized in a Tsa1-dependent manner. Conversely, peroxiredoxin hyperoxidation interrupts the H 2 O 2 signal and drives Msn2 oscillations by superimposing on PKA feedback regulation. Our data identify a mechanism by which light could be sensed in all cells lacking dedicated photoreceptors. In particular, the use of H 2 O 2 as a second messenger in signalling is common to Msn2 oscillations and to light-induced entrainment of circadian rhythms and suggests conserved roles for peroxiredoxins in endogenous rhythms.