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Ubiquitin Family Modifiers and the Proteasome

Volume 832 of the series Methods in Molecular Biology pp 363-371

Date:

Using DNA Damage Sensitivity Phenotypes to Characterize Mutations Affecting Proteasome Function

  • Benoît Le TallecAffiliated withCEA, iBiTecS, SBIGeM, Laboratoire du métabolisme de l’ADN et réponses aux génotoxiquesCentre de Recherche, Institut Curie
  • , Anne PeyrocheAffiliated withCEA, iBiTecS, SBIGeM, Laboratoire du métabolisme de l’ADN et réponses aux génotoxiques Email author 

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Abstract

Most mutants affected either in the proteasome biogenesis or function accumulate polyubiquitylated proteins and display growth defects at 37°C or in the presence of canavanine, an arginine analog that impairs protein synthesis. We uncovered a new striking phenotype related to DNA damage for some proteasome mutants: mutant strains grew better than the wild type in the presence of specific genotoxic agents (4NQO, Cpt, and MMS). Hyperresistance to 4NQO or Cpt is a new sensitive tool to detect proteasomal defects. Here, we describe simple methods that can be used to show and quantitatively measure this phenotype in budding yeast.

Key words

4-Nitroquinoline 1-oxide Camptothecin Hyperresistance DNA damage sensitivity Quantitative spotting