The dTAG System for Immediate and Target-Specific Protein Degradation

Nat Chem Biol. 2018 May;14(5):431-441. doi: 10.1038/s41589-018-0021-8. Epub 2018 Mar 26.

Abstract

Dissection of complex biological systems requires target-specific control of the function or abundance of proteins. Genetic perturbations are limited by off-target effects, multicomponent complexity, and irreversibility. Most limiting is the requisite delay between modulation to experimental measurement. To enable the immediate and selective control of single protein abundance, we created a chemical biology system that leverages the potency of cell-permeable heterobifunctional degraders. The dTAG system pairs a novel degrader of FKBP12F36V with expression of FKBP12F36V in-frame with a protein of interest. By transgene expression or CRISPR-mediated locus-specific knock-in, we exemplify a generalizable strategy to study the immediate consequence of protein loss. Using dTAG, we observe an unexpected superior antiproliferative effect of pan-BET bromodomain degradation over selective BRD4 degradation, characterize immediate effects of KRASG12V loss on proteomic signaling, and demonstrate rapid degradation in vivo. This technology platform will confer kinetic resolution to biological investigation and provide target validation in the context of drug discovery.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Animals
  • CRISPR-Cas Systems*
  • Cell Cycle Proteins
  • Cell Proliferation
  • Cytoplasm / metabolism
  • Dimerization
  • Gene Knock-In Techniques
  • HEK293 Cells
  • Homeostasis
  • Humans
  • Ligands
  • Mice
  • Mutation
  • NIH 3T3 Cells
  • Nuclear Proteins / chemistry*
  • Nuclear Proteins / genetics
  • Protein Binding
  • Protein Domains
  • Proteolysis
  • Proteomics
  • Proto-Oncogene Proteins p21(ras) / genetics*
  • Signal Transduction
  • Tacrolimus Binding Protein 1A / chemistry*
  • Transcription Factors / genetics*
  • Transgenes

Substances

  • BRD4 protein, human
  • Cell Cycle Proteins
  • KRAS protein, human
  • Ligands
  • Nuclear Proteins
  • Transcription Factors
  • Proto-Oncogene Proteins p21(ras)
  • Tacrolimus Binding Protein 1A