Grb10/Nedd4-mediated multiubiquitination of the insulin-like growth factor receptor regulates receptor internalization

J Cell Physiol. 2008 Aug;216(2):426-37. doi: 10.1002/jcp.21405.

Abstract

The adaptor protein Grb10 is an interacting partner of the IGF-I receptor (IGF-IR) and the insulin receptor (IR). Previous work from our laboratory has established the role of Grb10 as a negative regulator of IGF-IR-dependent cell proliferation. We have shown that Grb10 binds the E3 ubiquitin ligase Nedd4 and promotes IGF-I-stimulated ubiquitination, internalization, and degradation of the IGF-IR, thereby giving rise to long-term attenuation of signaling. Recent biochemical evidence suggests that tyrosine-kinase receptors (RTK) may not be polyubiquitinated but monoubiquitinated at multiple sites (multiubiquitinated). However, the type of ubiquitination of the IGF-IR is still not defined. Here we show that the Grb10/Nedd4 complex upon ligand stimulation mediates multiubiquitination of the IGF-IR, which is required for receptor internalization. Moreover, Nedd4 by promoting IGF-IR ubiquitination and internalization contributes with Grb10 to negatively regulate IGF-IR-dependent cell proliferation. We also demonstrate that the IGF-IR is internalized through clathrin-dependent and-independent pathways. Grb10 and Nedd4 remain associated with the IGF-IR in early endosomes and caveosomes, where they may participate in sorting internalized receptors. Grb10 and Nedd4, unlike the IGF-IR, which is targeted for lysosomal degradation are not degraded and likely directed into recycling endosomes. These results indicate that Grb10 and Nedd4 play a critical role in mediating IGF-IR down-regulation by promoting ligand-dependent multiubiquitination of the IGF-IR, which is required for receptor internalization and regulates mitogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Caveolin 1 / genetics
  • Caveolin 1 / metabolism
  • Cell Line
  • Cell Proliferation
  • Clathrin / genetics
  • Clathrin / metabolism
  • Endosomal Sorting Complexes Required for Transport
  • Endosomes / metabolism
  • GRB10 Adaptor Protein / genetics
  • GRB10 Adaptor Protein / metabolism*
  • Lysosomes / metabolism
  • Mice
  • Multiprotein Complexes / metabolism
  • Nedd4 Ubiquitin Protein Ligases
  • Polyubiquitin / metabolism*
  • RNA, Small Interfering / genetics
  • RNA, Small Interfering / metabolism
  • Rats
  • Receptor, IGF Type 1 / genetics
  • Receptor, IGF Type 1 / metabolism*
  • Receptor, Insulin / genetics
  • Receptor, Insulin / metabolism
  • Ubiquitin-Protein Ligases / genetics
  • Ubiquitin-Protein Ligases / metabolism*
  • Ubiquitination
  • Vesicular Transport Proteins / genetics
  • Vesicular Transport Proteins / metabolism

Substances

  • Caveolin 1
  • Clathrin
  • Endosomal Sorting Complexes Required for Transport
  • Multiprotein Complexes
  • RNA, Small Interfering
  • Vesicular Transport Proteins
  • early endosome antigen 1
  • Polyubiquitin
  • GRB10 Adaptor Protein
  • NEDD4L protein, rat
  • Nedd4 Ubiquitin Protein Ligases
  • Nedd4 protein, rat
  • Nedd4l protein, mouse
  • Ubiquitin-Protein Ligases
  • Receptor, IGF Type 1
  • Receptor, Insulin