pSILAC mass spectrometry reveals ZFP91 as IMiD-dependent substrate of the CRL4 CRBN ubiquitin ligase

Nat Commun. 2017 May 22;8:15398. doi: 10.1038/ncomms15398.

Abstract

Thalidomide and its derivatives lenalidomide and pomalidomide (IMiDs) are effective treatments of haematologic malignancies. It was shown that IMiDs impart gain-of-function properties to the CUL4-RBX1-DDB1-CRBN (CRL4CRBN) ubiquitin ligase that enable binding, ubiquitination and degradation of key therapeutic targets such as IKZF1, IKZF3 and CSNK1A1. While these substrates have been implicated as efficacy targets in multiple myeloma (MM) and 5q deletion associated myelodysplastic syndrome (del(5q)-MDS), other targets likely exist. Using a pulse-chase SILAC mass spectrometry-based proteomics approach, we demonstrate that lenalidomide induces the ubiquitination and degradation of ZFP91. We establish ZFP91 as a bona fide IMiD-dependent CRL4CRBN substrate and further show that ZFP91 harbours a zinc finger (ZnF) motif, related to the IKZF1/3 ZnF, critical for IMiD-dependent CRBN binding. These findings demonstrate that single time point pulse-chase SILAC mass spectrometry-based proteomics (pSILAC MS) is a sensitive approach for target identification of small molecules inducing selective protein degradation.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adaptor Proteins, Signal Transducing
  • Amino Acid Motifs
  • Animals
  • Antineoplastic Agents / pharmacology
  • Gene Deletion
  • HCT116 Cells
  • HEK293 Cells
  • Humans
  • Lenalidomide / pharmacology
  • Mass Spectrometry
  • Mice
  • Multiple Myeloma / metabolism
  • Myelodysplastic Syndromes / metabolism
  • Peptide Hydrolases / chemistry
  • Proteomics
  • Substrate Specificity
  • Ubiquitin / chemistry
  • Ubiquitin-Protein Ligases / chemistry*
  • Ubiquitination
  • Zinc Fingers

Substances

  • Adaptor Proteins, Signal Transducing
  • Antineoplastic Agents
  • CRBN protein, human
  • Ubiquitin
  • Ubiquitin-Protein Ligases
  • Peptide Hydrolases
  • CRL4 protein, human
  • ZFP91 protein, human
  • Lenalidomide