MDM2 Protein-mediated Ubiquitination of NUMB Protein IDENTIFICATION OF A SECOND PHYSIOLOGICAL SUBSTRATE OF MDM2 THAT EMPLOYS A DUAL-SITE DOCKING MECHANISM

M. Sczaniecka, K. Gladstone, S. Pettersson, L. McLaren, A. S. Huart, M. Wallace

Research output: Contribution to journalArticlepeer-review

Abstract

The E3 ubiquitin ligase, MDM2, uses a dual-site mechanism to ubiquitinate and degrade the tumor suppressor protein p53, involving interactions with the N-terminal hydrophobic pocket and the acidic domain of MDM2. The results presented here demonstrate that MDM2 also uses this same dual-site mechanism to bind to the cell fate determinant NUMB with both the N-terminal hydrophobic pocket and the acidic domain of MDM2 also involved in forming the interaction with NUMB. Furthermore, the acidic domain interactions are crucial for MDM2-mediated ubiquitination of NUMB. Contrary to p53, where two separate domains form the interface with MDM2, only one region within the phosphotyrosine binding domain of NUMB (amino acids 113-148) mediates binding to both these regions of MDM2. By binding to both domains on MDM2, NUMB disrupts the MDM2-p53 complex and MDM2-catalyzed ubiquitination of p53. Therefore, we have identified the mechanism NUMB uses to regulate the steady-state levels of the p53 in cells. By targeting the acidic domain of MDM2 using acid domain-binding ligands we can overcome MDM2-mediated ubiquitination and degradation of NUMB impacting on the stabilization of p53 in cells. Furthermore, delivery of MDM2 acid domain-binding ligands to cancer cells promotes p53-dependent growth arrest and the induction of apoptosis. This highlights the dual-site mechanism of MDM2 on another physiological substrate and identifies the acid domain as well as N terminus as a potential target for small molecules that inhibit MDM2.
Original languageEnglish
Pages (from-to)14052-14068
Number of pages17
JournalJournal of Biological Chemistry
Volume287
Issue number17
DOIs
Publication statusPublished - 2012

Keywords

  • APOPTOSIS
  • Binding Sites
  • Cell Line, Tumor
  • Cell Lineage
  • Cell Proliferation
  • Enzyme-Linked Immunosorbent Assay
  • Flow Cytometry
  • HUMANS
  • LIGANDS
  • Membrane Proteins
  • Nerve Tissue Proteins
  • Protein Binding
  • Protein Structure, Tertiary
  • Proto-Oncogene Proteins c-mdm2
  • Tumor Suppressor Protein p53
  • Ubiquitin
  • Ubiquitin-Protein Ligases
  • UBIQUITINATION

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