Regulation of redox homeostasis in the yeast <i>Saccharomyces cerevisiae</i>

Wheeler, Glen; Grant, Chris M.

doi:10.1111/j.0031-9317.2004.0193.x

Cited by 69 publications

(44 citation statements)

References 90 publications

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“…Article and publication date are available at www.molbiolcell.org/cgi/doi/10.1091/mbc. E04 -06 -0445. responsible for repairing oxidative damage to cytosolic proteins (for review, see Wheeler and Grant, 2004). The results reported here suggest that S. cerevisiae actin is susceptible to the same oxidative damage as red blood cell actin and that like ISC red blood cells, disulfide bond formation in actin is a major source of the physiological response and sensitivity to oxidative stress.…”

Section: Introductionmentioning

confidence: 74%

Old Yellow Enzyme Protects the Actin Cytoskeleton from Oxidative Stress

Haarer

Amberg

2004

MBoC

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Old Yellow Enzyme (OYE) has long served as a paradigm for the study of flavin-containing NADPH oxido-reductases and yet its physiological role has remained a mystery. A two-hybrid interaction between Oye2p and actin led us to investigate a possible function in the actin cytoskeleton. We found that oye deletion strains have an overly elaborate actin cytoskeleton that cannot be attributed to changes in actin concentration but likely reflect stabilization of actin filaments, resulting in excessive actin assembly. Cells expressing the actin mutant act1-123p, which has a weakened interaction with Oye2p, show comparable defects in actin organization to the oye deletion strain that can be suppressed by overexpression of Oye2p. Similarly, mutation of either conserved cysteine of the potential disulfide pair Cys285-Cys374 in actin completely suppresses the actin organization defect of the oye⌬ phenotype. Strains lacking Oye function are also sensitive to oxidative stress as induced by H 2 O 2 , menadione, and diamide treatment. Mutation of either Cys285 or Cys374 of actin suppresses the sensitivity of oye⌬ strains to oxidative stress and in fact confers super-resistance to oxidative stress in otherwise wild-type strains. These results suggest that oxidative damage to actin, like that which has been observed in irreversibly sickled red blood cells, may be a general phenomenon and that OYE functions to control the redox state of actin thereby maintaining the proper plasticity of the actin cytoskeleton. In addition to uncovering a long sought biological function for Old Yellow Enzyme, these results establish that cellular sensitivity to oxidative stress can in part be directly attributed to a specific form (C285-C374 disulfide bond formation) of oxidative damage to actin.

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Section: Introductionmentioning

confidence: 74%

Old Yellow Enzyme Protects the Actin Cytoskeleton from Oxidative Stress

Haarer

Amberg

2004

MBoC

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“…Further evidence that thioredoxins form part of the cellular response to a reductive challenge came from the finding that TRX2 gene expression is induced in response to DTT treatment and does not depend on the presence of IRE1 (Trotter and Grant, 2002). Loss of thioredoxins is known to result in elevated glutathione levels, indicating a link between the thioredoxin system and glutathione metabolism in the cell (reviewed in Carmel-Harel and Storz, 2000;Wheeler and Grant, 2004). We proposed that the combination of exogenous DTT and high intracellular GSH results in an additive reductive stress that is toxic to thioredoxin mutants (Trotter and Grant, 2002).…”

Section: Introductionmentioning

confidence: 99%

The Thioredoxin System Protects Ribosomes against Stress-induced Aggregation

Rand

Grant

2006

MBoC

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We previously showed that thioredoxins are required for dithiothreitol (DTT) tolerance, suggesting they maintain redox homeostasis in response to both oxidative and reductive stress conditions. In this present study, we screened the complete set of viable deletion strains in Saccharomyces cerevisiae for sensitivity to DTT to identify cell functions involved in resistance to reductive stress. We identified 195 mutants, whose gene products are localized throughout the cell. DTT-sensitive mutants were distributed among most major biological processes, but they particularly affected gene expression, metabolism, and the secretory pathway. Strikingly, a mutant lacking TSA1, encoding a peroxiredoxin, showed a similar sensitivity to DTT as a thioredoxin mutant. Epistasis analysis indicated that thioredoxins function upstream of Tsa1 in providing tolerance to DTT. Our data show that the chaperone function of Tsa1, rather than its peroxidase function, is required for this activity. Cells lacking TSA1 were found to accumulate aggregated proteins, and this was exacerbated by exposure to DTT. Analysis of the protein aggregates revealed that they are predominantly composed of ribosomal proteins. Furthermore, aggregation was found to correlate with an inhibition of translation initiation. We propose that Tsa1 normally functions to chaperone misassembled ribosomal proteins, preventing the toxicity that arises from their aggregation. INTRODUCTIONCells originally evolved in a reducing environment. The accumulation of oxygen in the atmosphere then made possible the efficient energy generation process of respiration. Cells have therefore evolved to survive both oxidative and reductive conditions. Although much is now known regarding the damaging effects of oxidation, little is known regarding the molecular responses to a reducing environment, which is the subject of this present study.Glutathione is the most abundant low-molecular-weight thiol in eukaryotic cells and has been widely used as an indicator of the cellular redox state (Schafer and Buettner, 2001). The cytoplasm is generally very reducing; for example, the ratio of reduced glutathione (GSH) to oxidized glutathione (GSSG) in yeast is in the range of ϳ70 -190:1 (Grant et al., 1998;Garrido and Grant, 2002). In contrast, the endoplasmic reticulum is more oxidizing and the GSH:GSSG ratio has been measured in the range of 1:1-3:1 (Hwang et al., 1992;Bass et al., 2004). Disulfide bonds are essential for the folding and stability of proteins that are secreted or localized through the secretory pathway. In eukaryotic cells, disulfide bonds are formed in nascent proteins within the lumen of the endoplasmic reticulum (ER). Genetic evidence has identified a pathway for disulfide bond formation where oxidizing equivalents are transferred between thiol-containing proteins and secretory proteins (Frand et al., 2000;Gross et al., 2002). Thus, oxidative protein folding proceeds primarily by protein-protein-based relays. Glutathione provides a buffer against hyperoxidizing conditions in ...

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“…An example of such a pathway is present in the cytosol in which the poxvirus has a redox pathway for inserting disulfide bonds in the virion membrane (24); this pathway consists of a virally coded sulfhydryl oxidase and a thioredoxinlike protein. It is possible that the mitochondrial intermembrane space may possess a similar pathway with the resident protein Erv1p, a sulfhydryl oxidase, and a thioredoxin system (40,41).…”

Section: Discussionmentioning

confidence: 99%

The Role of Hot13p and Redox Chemistry in the Mitochondrial TIM22 Import Pathway

Curran¹,

Leuenberger²,

Leverich³

et al. 2004

Journal of Biological Chemistry

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The small Tim proteins in the mitochondrial intermembrane space participate in the TIM22 import pathway for assembly of the inner membrane. Assembly of the small TIM complexes requires the conserved "twin CX 3 C" motif that forms juxtapositional intramolecular disulfide bonds. Here we identify a new intermembrane space protein, Hot13p, as the first component of a pathway that mediates assembly of the small TIM complexes. The small Tim proteins require Hot13p for assembly into a 70-kDa complex in the intermembrane space. Once assembled the small TIM complexes escort hydrophobic inner membrane proteins en route to the TIM22 complex. The mechanism by which the small Tim proteins bind and release substrate is not understood, and we investigated the affect of oxidant/reductant treatment on the TIM22 import pathway. With in organello import studies, oxidizing agents arrest the ADP/ATP carrier (AAC) bound to the Tim9p-Tim10p complex in the intermembrane space; this productive intermediate can be chased into the inner membrane upon subsequent treatment with reductant. Moreover, AAC import is markedly decreased by oxidant treatment in ⌬hot13 mitochondria and improved when Hot13p is overexpressed, suggesting Hot13p may function to remodel the small TIM complexes during import. Together these results suggest that the small TIM complexes have a specialized assembly pathway in the intermembrane space and that the local redox state of the TIM complexes may mediate translocation of inner membrane proteins.Mitochondrial inner membrane biogenesis is an essential process in all eukaryotes. The Translocase of the Inner Membrane (TIM) 1 22 complex is dedicated to the insertion of nuclear-coded polytopic inner membrane proteins, including the mitochondrial carrier family and import components Tim22p and Tim23p (1-3). The 300-kDa membrane complex consists of a pore-forming subunit Tim22p, in addition to Tim18p and Tim54p, that plays an unknown function and a fraction of the small Tim proteins Tim9p, Tim10p, and Tim12p (1-3). Substrates are escorted from the Translocase of the Outer Membrane (TOM) complex to the TIM22 translocon by two distinct 70-kDa chaperone-like complexes in the intermembrane space: Tim9p-Tim10p and Tim8p-Tim13p (4 -7). Insertion into the inner membrane requires a membrane potential (⌬⌿), but the events by which a substrate is transported from the TOM complex to the inner membrane complex remain obscure. The carrier proteins and Tim23p are targeted to the Tom70p receptor on the outer membrane (7, 8) and then pass through the TOM complex as a loop (4,5,8). The small Tim proteins subsequently bind to hydrophobic domains in the substrates and chaperone them to the 300-kDa insertion complex (4, 5, 9). The substrate is transferred to the insertion complex and inserted into the inner membrane (10). Cryoelectron microscopy and reconstitution studies have shown that Tim22p forms a twin-pore translocase that is voltagegated (10, 11).The small Tim proteins contain the "twin CX 3 C" motif that is required for assembly of the...

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Regulation of redox homeostasis in the yeast Saccharomyces cerevisiae

Cited by 69 publications

References 90 publications

Old Yellow Enzyme Protects the Actin Cytoskeleton from Oxidative Stress

Old Yellow Enzyme Protects the Actin Cytoskeleton from Oxidative Stress

The Thioredoxin System Protects Ribosomes against Stress-induced Aggregation

The Role of Hot13p and Redox Chemistry in the Mitochondrial TIM22 Import Pathway

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