Journal Article

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

J.R. Merwin, D.J. Mustacich, E.G.D. Muller, G.D. Pearson and G.F. Merrill

in Carcinogenesis

Volume 23, issue 10, pages 1609-1616
Published in print October 2002 | ISSN: 0143-3334
Published online October 2002 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/23.10.1609
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

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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 Δtrr1 yeast. The Δtrr1 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 Δtrr1 yeast. About 18% of glutathione was in the GSSG form in wild-type yeast and increased to 32% in Δtrr1 yeast. The Δtrr1 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 Δtrr1 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 Δglr1 mutation, even though the mutation was known to result in glutathione oxidation. In summary, the results show that, although glutathione becomes more oxidized in Δtrr1 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.

Keywords: DIT, dithiothreitol; IAA, iodoacetate; IAM, iodoacetamide; PDTC, pyrrolidine dithiocarbamate; PEMSA, protein electrophoretic mobility shift assay

Journal Article.  6378 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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