Focal adhesion kinase (FAK) binds RET kinase via its FERM domain, priming a direct and reciprocal RET-FAK transactivation mechanism

J Biol Chem. 2011 May 13;286(19):17292-302. doi: 10.1074/jbc.M110.168500. Epub 2011 Mar 22.

Abstract

Whether RET is able to directly phosphorylate and activate downstream targets independently of the binding of proteins that contain Src homology 2 or phosphotyrosine binding domains and whether mechanisms in trans by cytoplasmic kinases can modulate RET function and signaling remain largely unexplored. In this study, oligopeptide arrays were used to screen substrates directly phosphorylated by purified recombinant wild-type and oncogenic RET kinase domain in the presence or absence of small molecule inhibitors. The results of the peptide array were validated by enzyme kinetics, in vitro kinase, and cell-based experiments. The identification of focal adhesion kinase (FAK) as a direct substrate for RET kinase revealed (i) a RET-FAK transactivation mechanism consisting of direct phosphorylation of FAK Tyr-576/577 by RET and a reciprocal phosphorylation of RET by FAK, which crucially is able to rescue the kinase-impaired RET K758M mutant and (ii) that FAK binds RET via its FERM domain. Interestingly, this interaction is abolished upon RET phosphorylation, indicating that RET binding to the FERM domain of FAK is a priming step for RET-FAK transactivation. Finally, our data indicate that FAK inhibitors could be used as potential therapeutic agents for patients with multiple endocrine neoplasia type 2 tumors because both, treatment with the FAK kinase inhibitor NVP-TAE226 and FAK down-regulation by siRNA reduced RET phosphorylation and signaling as well as the proliferation and survival of tumor and transfected cell lines expressing oncogenic RET.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Antineoplastic Agents / pharmacology
  • Cell Proliferation
  • Focal Adhesion Protein-Tyrosine Kinases / genetics
  • Focal Adhesion Protein-Tyrosine Kinases / metabolism*
  • Gene Expression Regulation, Enzymologic*
  • Glutathione Transferase / metabolism
  • Humans
  • Kinetics
  • Oligopeptides / chemistry
  • Phenotype
  • Phosphorylation
  • Protein Interaction Mapping
  • Protein Structure, Tertiary
  • Proto-Oncogene Proteins c-ret / genetics
  • Proto-Oncogene Proteins c-ret / metabolism*
  • Signal Transduction
  • Transcriptional Activation*

Substances

  • Antineoplastic Agents
  • Oligopeptides
  • Glutathione Transferase
  • Proto-Oncogene Proteins c-ret
  • Focal Adhesion Protein-Tyrosine Kinases