Subquinocin, a small molecule inhibitor of CYLD and USP-family deubiquitinating enzymes, promotes NF-κB signaling

Biochem Biophys Res Commun. 2020 Mar 26;524(1):1-7. doi: 10.1016/j.bbrc.2019.12.049. Epub 2019 Dec 30.


The tumor suppressor CYLD negatively regulates polyubiquitination-dependent cellular signaling such as nuclear factor (NF)-κB signaling. In addition to CYLD, multiple deubiquitinating enzymes (DUBs) are also involved in the regulation of this signaling pathway, and distinct role of CYLD is yet to be clarified. Here, we identified a small chemical named Subquinocin that inhibited the DUB activity of recombinant CYLD using a wheat cell-free protein synthesis and an AlphaScreen technology. In cells, Subquinocin increased the polyubiquitination of NEMO and RIP1 and enhanced NF-κB activation. Modeling and mutation analyses indicated that Subquinocin interacted with Y940 in CYLD, which locates close to catalytic center of CYLD, and is conserved among the USP-family DUBs. Further biochemical evaluation revealed that Subquinocin inhibited USP-family DUBs, but not other family DUBs including OTU. Although Subquinocin showed a broad specificity toward USP-family DUBs, the inhibitory effect of Subquinocin on NF-κB signaling was negligible in CYLD-KO cells, indicating that CYLD is a major target of Subquinocin on the suppression of NF-κB signaling. In conclusion, Subquinocin identified here is a useful tool to analyze the signal transduction mediated by USP-family DUBs.

Keywords: CYLD; Deubiquitinating enzyme; NF-κB signaling; Small molecule inhibitor; Ubiquitin.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Antineoplastic Agents / chemistry*
  • Antineoplastic Agents / pharmacology
  • Cell Line, Tumor
  • Deubiquitinating Enzyme CYLD / antagonists & inhibitors*
  • Drug Screening Assays, Antitumor
  • Enzyme Inhibitors / chemistry*
  • Enzyme Inhibitors / pharmacology
  • Gene Expression Regulation / drug effects
  • Genes, Tumor Suppressor / drug effects
  • Glutathione Transferase / metabolism
  • Humans
  • Molecular Docking Simulation
  • Mutation
  • NF-kappa B / metabolism*
  • Nuclear Pore Complex Proteins / metabolism
  • Protein Binding
  • Protein Conformation
  • RNA-Binding Proteins / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / metabolism
  • Signal Transduction
  • Tumor Suppressor Proteins / genetics
  • Tumor Suppressor Proteins / metabolism
  • Ubiquitin-Specific Proteases / metabolism
  • Ubiquitination / drug effects


  • AGFG1 protein, human
  • Antineoplastic Agents
  • Enzyme Inhibitors
  • NF-kappa B
  • Nuclear Pore Complex Proteins
  • RNA-Binding Proteins
  • Recombinant Proteins
  • Tumor Suppressor Proteins
  • Glutathione Transferase
  • Deubiquitinating Enzyme CYLD
  • Ubiquitin-Specific Proteases