Reporter gene transactivation by human p53 is inhibited in thioredoxin reductase null yeast by a mechanism associated with thioredoxin oxidation and independent of changes in the redox state of glutathione.
Reporter gene transactivation by human p53 is compromised in S. cerevisiae lacking the TRR1 gene encoding thioredoxin reductase. The basis for p53 inhibition was investigated by measuring the redox state of thioredoxin and glutathione in wild-type and Deltatrr1 yeast. The Deltatrr1 mutation affected the redox state of both molecules. About 34% of thioredoxin was in the disulfide form in wild-type yeast and increased to 70% in Deltatrr1 yeast. About 18% of glutathione was in the GSSG form in wild-type yeast and increased to 32% in Deltatrr1 yeast. The Deltatrr1 mutation also resulted in a 2.9-fold increase in total glutathione per mg extract protein. Highcopy expression of the GLR1 gene encoding glutathione reductase in Deltatrr1 yeast restored the GSSG:GSH ratio to wild-type levels, but did not restore p53 activity. Also, p53 activity was shown to be unaffected by a Deltaglr1 mutation, even though the mutation was known to result in glutathione oxidation. In summary, the results show that, although glutathione becomes more oxidized in Deltatrr1 cells, glutathione oxidation is neither sufficient nor necessary for p53 inhibition. The results indicate that p53 activity has a specific requirement for an intact thioredoxin system, rather than a general dependence on the intracellular reducing environment.
Merwin
J R
JR
Molecular and Cellular Biology Program, Oregon State University, Corvallis, Oregon 97331, USA.
Mustacich
D J
DJ
Muller
E G D
EG
Pearson
G D
GD
Merrill
G F
GF
D004220:Disulfides / Q000378:metabolism; D017353:Gene Deletion; D016158:Genes, p53; D005978:Glutathione / Q000378:metabolism*; D019803:Glutathione Disulfide / Q000378:metabolism; D006801:Humans; D010084:Oxidation-Reduction; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013880:Thioredoxin-Disulfide Reductase / Q000235:genetics*; D013879:Thioredoxins / Q000378:metabolism*; D015533:Transcriptional Activation / Q000502:physiology*; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism*
D004220:Disulfides; D013438:Sulfhydryl Compounds; D016159:Tumor Suppressor Protein p53; D013879:Thioredoxins; D013880:Thioredoxin-Disulfide Reductase; D005978:Glutathione; D019803:Glutathione Disulfide