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Thioredoxin-related Protein 32 is an arsenite-regulated Thiol Reductase of the proteasome 19 S particle.

Perturbation of the cytoplasmic protein folding environment by exposure to oxidative stress-inducing As(III)-containing compounds challenges the ubiquitin-proteasome system. Here we report on mass spectrometric analysis of As(III)-induced changes in the proteasome's composition in samples prepared by stable isotope labeling with amino acids in cell culture, using mammalian cells in which TRP32 (thioredoxin-related protein of 32 kDa; also referred to as TXNL1) was identified as a novel subunit of the 26 S proteasome. Quantitative genetic interaction mapping, using the epistatic miniarray profiling approach, identified a functional connection between TRP32 and the proteasome. Deletion of txl1, the Schizosaccharomyces pombe homolog of TRP32, results in a slow growth phenotype when combined with deletion of cut8, a gene required for normal proteasome localization. Deletion analysis in vivo, chemical cross-linking, and manipulation of the ATP concentration in vitro during proteasome immunopurification revealed that the C-terminal domain of mammalian TRP32 binds the 19 S regulatory particle in proximity to the proteasome substrate binding site. Thiol modification with polyethylene glycol-maleimide showed disulfide bond formation at the active site of TRP32 in cells exposed to As(III). Pulse-chase labeling showed that TRP32 is a stable protein whose half-life of >6 h is surprisingly reduced to 1 h upon exposure of cells to As(III). These findings reveal a previously undescribed thiol reductase at the proteasome's regulatory particle.

Pubmed ID: 19349280 RIS Download

Mesh terms: Animals | Arsenites | Cell Line | Disulfides | Humans | Isotope Labeling | Mice | Mutagenesis, Insertional | Oxidation-Reduction | Oxidative Stress | Proteasome Endopeptidase Complex | Protein Binding | Protein Disulfide Reductase (Glutathione) | Protein Processing, Post-Translational | Protein Structure, Tertiary | Schizosaccharomyces | Thioredoxins

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Associated grants

  • Agency: NIEHS NIH HHS, Id: F32-ES014775
  • Agency: NIGMS NIH HHS, Id: R01-GM084279
  • Agency: NINDS NIH HHS, Id: P30 NS050276
  • Agency: NINDS NIH HHS, Id: F32-NS050901
  • Agency: NIGMS NIH HHS, Id: R01 GM084448
  • Agency: NIEHS NIH HHS, Id: R01-ES08681
  • Agency: NINDS NIH HHS, Id: P30-NS050276
  • Agency: NIGMS NIH HHS, Id: R01 GM084279
  • Agency: PHS HHS, Id: MC-IRG203123

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