Regulation of Nedd4-2 self-ubiquitination and stability by a PY motif located within its HECT-domain

Biochem J. 2008 Oct 1;415(1):155-63. doi: 10.1042/BJ20071708.


Ubiquitin ligases play a pivotal role in substrate recognition and ubiquitin transfer, yet little is known about the regulation of their catalytic activity. Nedd4 (neural-precursor-cell-expressed, developmentally down-regulated 4)-2 is an E3 ubiquitin ligase composed of a C2 domain, four WW domains (protein-protein interaction domains containing two conserved tryptophan residues) that bind PY motifs (L/PPXY) and a ubiquitin ligase HECT (homologous with E6-associated protein C-terminus) domain. In the present paper we show that the WW domains of Nedd4-2 bind (weakly) to a PY motif (LPXY) located within its own HECT domain and inhibit auto-ubiquitination. Pulse-chase experiments demonstrated that mutation of the HECT PY-motif decreases the stability of Nedd4-2, suggesting that it is involved in stabilization of this E3 ligase. Interestingly, the HECT PY-motif mutation does not affect ubiquitination or down-regulation of a known Nedd4-2 substrate, ENaC (epithelial sodium channel). ENaC ubiquitination, in turn, appears to promote Nedd4-2 self-ubiquitination. These results support a model in which the inter- or intra-molecular WW-domain-HECT PY-motif interaction stabilizes Nedd4-2 by preventing self-ubiquitination. Substrate binding disrupts this interaction, allowing self-ubiquitination of Nedd4-2 and subsequent degradation, resulting in down-regulation of Nedd4-2 once it has ubiquitinated its target. These findings also point to a novel mechanism employed by a ubiquitin ligase to regulate itself differentially compared with substrate ubiquitination and stability.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Motifs / physiology
  • Amino Acid Sequence
  • Animals
  • Drug Stability
  • Endosomal Sorting Complexes Required for Transport
  • Epithelial Sodium Channels / physiology
  • Humans
  • Models, Molecular
  • Molecular Sequence Data
  • Nedd4 Ubiquitin Protein Ligases
  • Oocytes / physiology
  • Protein Structure, Tertiary
  • Sequence Alignment
  • Ubiquitin-Protein Ligases / physiology*
  • Ubiquitination / physiology*
  • Xenopus Proteins
  • Xenopus laevis


  • Endosomal Sorting Complexes Required for Transport
  • Epithelial Sodium Channels
  • Xenopus Proteins
  • Nedd4 Ubiquitin Protein Ligases
  • Nedd4 protein, Xenopus
  • Nedd4 protein, human
  • Nedd4L protein, human
  • nedd4l protein, Xenopus
  • Ubiquitin-Protein Ligases