Characterizing ErbB-2-Mediated Tyrosine Phosphorylation and Activation of Plexins

Methods Mol Biol. 2017:1493:129-146. doi: 10.1007/978-1-4939-6448-2_9.

Abstract

Plexins comprise a family of transmembrane receptors for semaphorins. Plexins of the B- and D-subfamily interact with the receptor tyrosine kinase ErbB-2, and this interaction has been shown to be functionally relevant for various biological processes including tumor metastasis and bone formation. Binding of semaphorins to B- and D-subfamily plexins results in the activation of ErbB-2, which in turn phosphorylates these plexins. This phosphorylation triggers the activation of the small GTPases RhoA and RhoC downstream of B-subfamily plexins. Here we describe a methodology that allows the analysis of ErbB-2-mediated plexin phosphorylation and signaling.

Keywords: ErbB-2; Immunoblot; Immunoprecipitation; Phosphorylation; Plexin; Pulldown; RhoA; RhoC; Semaphorin; Signaling.

MeSH terms

  • Blotting, Western
  • Cell Adhesion Molecules / metabolism*
  • Electrophoresis, Polyacrylamide Gel
  • HEK293 Cells
  • Humans
  • MCF-7 Cells
  • Nerve Tissue Proteins / metabolism*
  • Phosphorylation
  • Receptor, ErbB-2 / metabolism*
  • Recombinant Fusion Proteins / metabolism
  • Tyrosine / metabolism*
  • rhoA GTP-Binding Protein / metabolism

Substances

  • Cell Adhesion Molecules
  • Nerve Tissue Proteins
  • Recombinant Fusion Proteins
  • plexin
  • Tyrosine
  • Receptor, ErbB-2
  • rhoA GTP-Binding Protein