Glutathione as an endogenous sulphur source in the yeast Saccharomyces cerevisiae

Abstract

Glutathione-deficient mutants (@A) of the yeast Saccharomyces cerevisiae, impaired in the first step of glutathione (GSH) biosynthesis were studied with respect to the regulation of enzymes involved in GSH catabolism and cysteine biosynthesis. Striking differences were observed in the content of the sulphur amino acids when gshA mutants were compared to wild-type strains growing on the same minimal medium. Furthermore, all mutants examined showed a derepression of y-glutamyltranspeptidase (y-GT), the enzyme initiating GSH degradation. However, y-cystathionase and cysteine synthase were unaffected by the GSH deficiency as long as the nutrient sulphate source was not exhausted. The results suggest that the mutants are probably not impaired in the sulphate assimilation pathway, but that the y-glutamyl cycle could play a leading role in the regulation of the sulphur fluxes. Studies of enzyme regulation showed that the derepression of y-GT observed in thegshA strains was most probably due to an alteration of the thiol status. The effectors governing the biosynthesis of cysteine synthase and ycystathionase seemed different from those playing a role in y-GT regulation and it was only under conditions of total sulphate deprivation that all these enzymes were derepressed. As a consequence the endogenous pool of GSH was used in the synthesis of cysteine. GSH might, therefore, fulfil the role of a storage compound.

Mutations in the CYS4 Gene Provide Evidence for Regulation of the Yeast Vacuolar H+-ATPase by Oxidation and Reduction in Vivo

“…Most of the glutathione in yeast cells is in the reduced (GSH) form; the thiol:disulfide (i.e. GSH:GSSG) balance is between 50 and 60, similar to other eukaryotic cells (46,47). Since cys4 mutants have been shown to have reduced cellular glutathione concentration (46), and our results (above) indicate that the V-ATPase is expressed at levels similar to wild type in cys4-⌬1 cells, we reasoned that the vma phenotypes of cys4 mutants may result from accumulation of the V-ATPase in an oxidized, inactive form in the less reducing environment resulting from low cellular GSH levels.…”

“…Chaudhuri et al (42) have reported that supplementation of Gsh Ϫ mutants of S. pombe with reduced glutathione (up to 0.25 mM) restores a Gsh ϩ growth phenotype but not pigmentation in adenine-limiting medium. The emerging theme from studies of GSH metabolism is that there is a minimum critical level of GSH required for growth and protein synthesis which is lower than that required for the other functions of GSH in metabolism, detoxification, and transport (47).…”

“…46). Since GSH represents more than 95% of the non-protein thiol pool in wild type yeast cells (47), GSH-deficient strains should have a much less reducing cytosol than wild type cells.…”

Physiological analysis of mutants of Saccharomyces cerevisiae impaired in sulphate assimilation

“…cerevisiae can use inorganic sulphur sources such as sulphate, sulphite or thiosulphate, as well as organic sulphur sources such as homocysteine, cysteine, methionine and S-adenosylmethionine. S. cereuisiae has also been shown to grow on glutathione (Elskens et al, 1991).…”

Single yeast cells vary in transcription activity not in delay time after a metabolic shift

“…This reduction is not enough to sustain measurable growth as the sulphur requirement of yeast is E90 nmol per mg dry weight 37 , indicating rapid sulphur limitation. Even if GSH reserves, with GSH concentrations about 10-fold higher than methionine concentrations 38,39 , would be used as a sulphur source this would not lead to significant growth. So, cells need to activate the sulphate assimilation pathway to synthesize methionine before they can continue growing.…”