The auxin-inducible degron 2 technology provides sharp degradation control in yeast, mammalian cells, and mice

Nat Commun. 2020 Nov 11;11(1):5701. doi: 10.1038/s41467-020-19532-z.


Protein knockdown using the auxin-inducible degron (AID) technology is useful to study protein function in living cells because it induces rapid depletion, which makes it possible to observe an immediate phenotype. However, the current AID system has two major drawbacks: leaky degradation and the requirement for a high dose of auxin. These negative features make it difficult to control precisely the expression level of a protein of interest in living cells and to apply this method to mice. Here, we overcome these problems by taking advantage of a bump-and-hole approach to establish the AID version 2 (AID2) system. AID2, which employs an OsTIR1(F74G) mutant and a ligand, 5-Ph-IAA, shows no detectable leaky degradation, requires a 670-times lower ligand concentration, and achieves even quicker degradation than the conventional AID. We demonstrate successful generation of human cell mutants for genes that were previously difficult to deal with, and show that AID2 achieves rapid target depletion not only in yeast and mammalian cells, but also in mice.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Cycle Proteins / genetics
  • Cell Cycle Proteins / metabolism
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism
  • Female
  • HCT116 Cells
  • Hippocampus / cytology
  • Humans
  • Indoleacetic Acids / pharmacology
  • Male
  • Mice, Inbred BALB C
  • Mice, Inbred C57BL
  • Mice, Transgenic
  • Minichromosome Maintenance Proteins / genetics
  • Minichromosome Maintenance Proteins / metabolism
  • Mutation
  • Neurons / drug effects
  • Neurons / metabolism
  • Nuclear Proteins / genetics
  • Nuclear Proteins / metabolism
  • Oryza / genetics
  • Plant Proteins / genetics
  • Plant Proteins / metabolism
  • Proteolysis / drug effects*
  • Proteomics / methods*
  • Recombinant Fusion Proteins / genetics
  • Recombinant Fusion Proteins / metabolism*
  • Saccharomyces cerevisiae Proteins / genetics
  • Saccharomyces cerevisiae Proteins / metabolism
  • Xenograft Model Antitumor Assays


  • CDC45 protein, S cerevisiae
  • Cell Cycle Proteins
  • DNA-Binding Proteins
  • Indoleacetic Acids
  • MCM10 protein, S cerevisiae
  • Nuclear Proteins
  • Plant Proteins
  • Recombinant Fusion Proteins
  • Saccharomyces cerevisiae Proteins
  • Sld3 protein, S cerevisiae
  • indoleacetic acid
  • Minichromosome Maintenance Proteins