The Yeast Copper/Zinc Superoxide Dismutase and the Pentose Phosphate Pathway Play Overlapping Roles in Oxidative Stress Protection

Abstract

In Saccharomyces cerevisiae, loss of cytosolic superoxide dismutase (Sod1) results in several air-dependent mutant phenotypes, including methionine auxotrophy and oxygen sensitivity. Here we report that these two sod1⌬ phenotypes were specifically suppressed by elevated expression of the TKL1 gene, encoding transketolase of the pentose phosphate pathway. The apparent connection between Sod1 and the pentose phosphate pathway prompted an investigation of mutants defective in glucose-6-phosphate dehydrogenase (Zwf1), which catalyzes the rate-limiting NADPH-producing step of this pathway. We confirmed that zwf1⌬ mutants are methionine auxotrophs and report that they also are oxygen-sensitive. We determined that a functional ZWF1 gene product was required for TKL1 to suppress sod1⌬, leading us to propose that increased flux through the oxidative reactions of the pentose phosphate pathway can rescue sod1 methionine auxotrophy. To better understand this methionine growth requirement, we examined the sulfur compound requirements of sod1⌬ and zwf1⌬ mutants, and noted that these mutants exhibit the same apparent defect in sulfur assimilation. Our studies suggest that this defect results from the impaired redox status of aerobically grown sod1 and zwf1 mutants, implicating Sod1 and the pentose phosphate pathway as being critical for maintenance of the cellular redox state.Aerobic organisms are constantly exposed to potentially harmful reactive oxygen species that are generated as byproducts of cellular metabolism. These oxygen free radicals have the potential to damage critical cellular components such as DNA, proteins, and lipids (see Refs. 1 and 2 for review). However, aerobic organisms have evolved with both enzymatic and non-enzymatic oxidant defense mechanisms that are designed for protection against such assaults. Non-enzymatic defenses include the tripeptide glutathione (GSH) and the small peptide thioredoxin (Trx), while enzymatic defenses include catalases, peroxidases, and superoxide dismutases (Ref.

Mechanisms for activating Cu- and Zn-containing superoxide dismutase in the absence of the CCS Cu chaperone

“…Plasmid transformed cells were grown on SD synthetic selecting medium that was supplemented with 1 mM glutathione for cultivation of gsh1⌬ mutants. Yeast strains KS107 (sod1⌬:TRP1) (20), LS101 (sod1⌬::TRP1 lys7⌬::URA3) (4), PS131 (lys7⌬::TRP1), PS132 (lys7⌬::TRP1 gsh1⌬::LEU2), and MC107 (sod1⌬::TRP1 gsh1⌬::LEU2) were all derived from the parental strain EG103 (MAT␣ leu 2-3, 112 his3⌬1 trp1-289 ura3-52). Strains 2737 (glr1⌬::kanMX4) and 614 (lys7⌬::kanMX4) are derivatives of BY4741 (MATa his3⌬1 leu2⌬0 met15⌬0 ura3⌬0; Research Genetics, Huntsville, AL).…”

Activation of CuZn Superoxide Dismutases from Caenorhabditis elegans Does Not Require the Copper Chaperone CCS

“…Strains and Culture Conditions-Yeast strains used in this study were derived from EG103 (MAT␣, leu2-3,112, his3⌬1, trp1-289, ura3-52) (35) and include KS107 (sod1⌬) (36), LS101 (sod1⌬ ccs1⌬) (10), MC107 (sod1⌬ gsh1⌬) (30), and LJ349 (sod1⌬ ccs1⌬ gsh1⌬). Disruption of CCS1 was generated with plasmid pPS003 as described previously (37).…”

Characterization of a CuZn superoxide dismutase gene in the arbuscular mycorrhizal fungus Glomus intraradices

“…Phenotypes of the SOD1 deficiency in aerobically grown yeast include retarded growth rate and auxotrophy for cysteine, lysine and methionine (Liu et al 1992), since the synthetic pathways for these amino acids contain steps that are hypersensitive to reactive oxygen (Liu et al 1992; Slekar et al 1996). To determine whether the GintSOD1 gene product could restore cysteine, lysine and methionine synthesis to aerobically grown SOD1-deficient yeast, the full-length cDNA was cloned into the pYES2 vector and expressed in the mutant strain (Fig.…”