Endoplasmic reticulum (ER) stress–induced reactive oxygen species (ROS) are detrimental for the fitness of a thioredoxin reductase mutant

Paraskevi Kritsiligkou,Jonathan D Rand,Alan J Weids,Ximeng Wang,Chris J Kershaw,Chris M Grant

doi:10.1074/jbc.ra118.001824

Paraskevi Kritsiligkou, Jonathan D Rand + Show 4 more

Open Access

https://doi.org/10.1074/jbc.ra118.001824

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Abstract

The unfolded protein response (UPR) is constitutively active in yeast thioredoxin reductase mutants, suggesting a link between cytoplasmic thiol redox control and endoplasmic reticulum (ER) oxidative protein folding. The unique oxidative environment of the ER lumen requires tight regulatory control, and we show that the active UPR depends on the presence of oxidized thioredoxins rather than arising because of a loss of thioredoxin function. Preventing activation of the UPR by deletion of HAC1, encoding the UPR transcription factor, rescues a number of thioredoxin reductase mutant phenotypes, including slow growth, shortened longevity, and oxidation of the cytoplasmic GSH pool. This is because the constitutive UPR in a thioredoxin reductase mutant results in the generation of hydrogen peroxide. The oxidation of thioredoxins in a thioredoxin reductase mutant requires aerobic metabolism and the presence of the Tsa1 and Tsa2 peroxiredoxins, indicating that a complete cytoplasmic thioredoxin system is crucial for maintaining ER redox homeostasis.

Highlights

The unfolded protein response (UPR) is constitutively active in yeast thioredoxin reductase mutants, suggesting a link between cytoplasmic thiol redox control and endoplasmic reticulum (ER) oxidative protein folding
This difference in phenotype arises because of differences in the regulation of TRX1 and TRX2 gene expression, as overexpression of either isoform increases resistance to oxidative stress [8]. In agreement with this idea, TRX2 is a target of the Yap1 transcription factor, which is the major regulator of oxidative stress–related genes in Saccharomyces cerevisiae, and its expression is up-regulated in a Yap1-dependent manner following ROS3 exposure [7]
We show that the UPR generates ROS in a trr1 mutant and that ROS generation accounts for the well-known phenotypes that are displayed by thioredoxin reductase mutants, including slow growth and shortened chronological life span

Summary

To whom correspondence should be addressed

The University of Manchester, Faculty of Biology, Medicine, and Health, Division of Molecular and Cellular Function, The Michael Smith Building, Oxford Road, Manchester M13 9PT, United Kingdom. Trx appears to be more important as an antioxidant because strains deleted for TRX2 are sensitive to hydrogen peroxide [7], whereas strains deleted for TRX1 are unaffected in oxidant sensitivity [8] This difference in phenotype arises because of differences in the regulation of TRX1 and TRX2 gene expression, as overexpression of either isoform increases resistance to oxidative stress [8]. Cysteine residues react relatively slowly with H2O2, but reactivity can be significantly enhanced by their ionization state, which depends on the local protein environment This means that some redox-regulated proteins are directly oxidized by ROS, it is thought that most regulatory thiol oxidation is mediated by protein catalysts such as peroxiredoxins [11, 12]. UPR-generated ROS activate the Yap transcription factor, and Yap is required for ROS tolerance under conditions of ER stress

Results

Discussion

Experimental procedures