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PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.
Journal article

PIKES Analysis Reveals Response to Degraders and Key Regulatory Mechanisms of the CRL4 Network.

  • Reichermeier KM Division of Biology and Biological Engineering, California Institute of Technology, 1200 E California Blvd., Pasadena, CA 91125, USA; Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA. Electronic address: kumire@caltech.edu.
  • Straube R Max Plank Institute for Dynamics of Complex Technical Systems, Sandtorstr. 1, 39106 Magdeburg, Germany; Bristol-Myers Squibb, 3551 Lawrenceville Princeton Rd, Lawrence Township, NJ 08648, USA.
  • Reitsma JM Division of Biology and Biological Engineering, California Institute of Technology, 1200 E California Blvd., Pasadena, CA 91125, USA; Abbvie, 1 N Waukegan Rd, North Chicago, IL 60064, USA.
  • Sweredoski MJ Division of Biology and Biological Engineering, California Institute of Technology, 1200 E California Blvd., Pasadena, CA 91125, USA.
  • Rose CM Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA.
  • Moradian A Division of Biology and Biological Engineering, California Institute of Technology, 1200 E California Blvd., Pasadena, CA 91125, USA.
  • den Besten W Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA; Amgen Research, Amgen, One Amgen Center Drive, 29MB, Thousand Oaks, CA 91320, USA.
  • Hinkle T Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA.
  • Verschueren E Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA.
  • Petzold G Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel, Switzerland.
  • Thomä NH Friedrich Miescher Institute for Biomedical Research, Maulbeerstrasse 66, 4058 Basel, Switzerland.
  • Wertz IE Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA.
  • Deshaies RJ Division of Biology and Biological Engineering, California Institute of Technology, 1200 E California Blvd., Pasadena, CA 91125, USA; Amgen Research, Amgen, One Amgen Center Drive, 29MB, Thousand Oaks, CA 91320, USA.
  • Kirkpatrick DS Genentech, 1 DNA Way, South San Francisco, 94080 CA, USA. Electronic address: donaldk@gene.com.
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  • 2020-01-25
Published in:
  • Molecular cell. - 2020
CRL4
DCAFs
Nedd8
PIKES
QconCAT
cullin-RING ligase
mass spectrometry
quantitative proteomics
targeted protein degradation
ubiquitin
Chromatography, High Pressure Liquid
Cullin Proteins
HEK293 Cells
Humans
Kinetics
Muscle Proteins
NEDD8 Protein
Protein Interaction Maps
Proteolysis
Proteomics
Signal Transduction
Spectrometry, Mass, Electrospray Ionization
Tandem Mass Spectrometry
Transcription Factors
Ubiquitin-Protein Ligases
English Co-opting Cullin4 RING ubiquitin ligases (CRL4s) to inducibly degrade pathogenic proteins is emerging as a promising therapeutic strategy. Despite intense efforts to rationally design degrader molecules that co-opt CRL4s, much about the organization and regulation of these ligases remains elusive. Here, we establish protein interaction kinetics and estimation of stoichiometries (PIKES) analysis, a systematic proteomic profiling platform that integrates cellular engineering, affinity purification, chemical stabilization, and quantitative mass spectrometry to investigate the dynamics of interchangeable multiprotein complexes. Using PIKES, we show that ligase assemblies of Cullin4 with individual substrate receptors differ in abundance by up to 200-fold and that Cand1/2 act as substrate receptor exchange factors. Furthermore, degrader molecules can induce the assembly of their cognate CRL4, and higher expression of the associated substrate receptor enhances degrader potency. Beyond the CRL4 network, we show how PIKES can reveal systems level biochemistry for cellular protein networks important to drug development.
Language
  • English
Open access status
closed
Identifiers
Persistent URL
https://sonar.ch/global/documents/299275
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