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Regulation of Mdm2 protein stability and the p53 response by NEDD4-1 E3 ligase

C Xu1, C D Fan1, X Wang1

  • 1Department of Pharmacology and Therapeutics, Roswell Park Cancer Institute, Buffalo, NY, USA.

Oncogene
|January 14, 2014
PubMed

Insights

The E3 ligase NEDD4-1 stabilizes the Mdm2 protein, a key regulator of tumor suppressor p53. This interaction impacts p53 activity and cellular responses to DNA damage, revealing NEDD4-1 as a novel component in the p53/Mdm2 feedback loop.

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Mdm2 is a crucial negative regulator of the tumor suppressor p53, acting as an E3 ligase that targets p53 for degradation.
  • Mdm2 protein stability is tightly controlled, involving mechanisms like autoubiquitination mediated by its RING domain.

Purpose of the Study:

  • To identify novel E3 ligases that regulate Mdm2 protein stability.
  • To elucidate the role of NEDD4-1 in the p53/Mdm2 regulatory feedback loop.

Main Methods:

  • Biochemical identification of E3 ligases using in vitro assays with an enzyme-dead Mdm2 mutant.
  • Analysis of Mdm2 ubiquitination and stability in Nedd4-1 knockout (KO) mouse embryonic fibroblasts (MEFs) and cell lines with NEDD4-1 knockdown.
  • Investigation of protein-protein interactions between NEDD4-1 and Mdm2 using cell-based assays.
  • Assessment of p53 levels, activity, and cellular responses (DNA damage response, cell cycle arrest) in the presence and absence of NEDD4-1.

Main Results:

  • NEDD4-1 was identified as an E3 ligase for Mdm2, promoting its ubiquitination.
  • NEDD4-1 catalyzes K63-type polyubiquitination of Mdm2, which competes with K48-type ubiquitination and stabilizes Mdm2 protein.
  • NEDD4-1 knockdown significantly reduces Mdm2 half-life and increases p53 levels and activity.
  • Loss of NEDD4-1 function in MEFs or cells leads to enhanced p53-mediated responses to DNA damage and growth inhibition.

Conclusions:

  • NEDD4-1 is a novel E3 ligase that stabilizes Mdm2 through K63-specific ubiquitination.
  • NEDD4-1 functions as a critical component of the p53/Mdm2 regulatory feedback loop, influencing p53 activity and cellular stress responses.
  • Targeting NEDD4-1 may offer a therapeutic strategy for enhancing tumor suppressor p53 activity.

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