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Polyubiquitin-sensor proteins reveal localization and linkage-type dependence of cellular ubiquitin signaling.

Polyubiquitin chain topology is thought to direct modified substrates to specific fates, but this function-topology relationship is poorly understood, as are the dynamics and subcellular locations of specific polyubiquitin signals. Experimental access to these questions has been limited because linkage-specific inhibitors and in vivo sensors have been unavailable. Here we present a general strategy to track linkage-specific polyubiquitin signals in yeast and mammalian cells, and to probe their functions. We designed several high-affinity Lys63 polyubiquitin-binding proteins and demonstrate their specificity in vitro and in cells. We apply these tools as competitive inhibitors to dissect the polyubiquitin-linkage dependence of NF-κB activation in several cell types, inferring the essential role of Lys63 polyubiquitin for signaling via the IL-1β and TNF-related weak inducer of apoptosis (TWEAK) but not TNF-α receptors. We anticipate live-cell imaging, proteomic and biochemical applications for these tools and extension of the design strategy to other polymeric ubiquitin-like protein modifications.

Pubmed ID: 22306808

Authors

  • Sims JJ
  • Scavone F
  • Cooper EM
  • Kane LA
  • Youle RJ
  • Boeke JD
  • Cohen RE

Journal

Nature methods

Publication Data

March 29, 2012

Associated Grants

  • Agency: NIGMS NIH HHS, Id: 1R01 GM097452
  • Agency: NCI NIH HHS, Id: P01 CA139980
  • Agency: NIGMS NIH HHS, Id: R01 GM097452
  • Agency: NIGMS NIH HHS, Id: R01 GM097452-01
  • Agency: NIGMS NIH HHS, Id: RC1 GM091424
  • Agency: NIGMS NIH HHS, Id: RC1 GM091424
  • Agency: NIGMS NIH HHS, Id: RC1 GM091424-01
  • Agency: NIGMS NIH HHS, Id: RC1 GM091424-02
  • Agency: NCRR NIH HHS, Id: RR020839
  • Agency: Intramural NIH HHS, Id:

Mesh Terms

  • Animals
  • Binding Sites
  • Humans
  • Molecular Probe Techniques
  • Protein Binding
  • Protein Interaction Mapping
  • Signal Transduction
  • Ubiquitin