Docking Protein FRS2 Links the Protein Tyrosine Kinase RET and Its Oncogenic Forms with the Mitogen-Activated Protein Kinase Signaling Cascade

Abstract

The receptor tyrosine kinase RET functions as the signal transducing receptor for the GDNF (for “glial cell-derived neurotrophic factors”) family of ligands. Mutations in the RET gene were implicated in Hirschsprung disease (HSCR), multiple endocrine neoplasia type 2 (MEN 2), and thyroid carcinomas. In this report we demonstrate that the docking protein FRS2 is tyrosine phosphorylated by ligand-stimulated and by constitutively activated oncogenic forms of RET. Complex formation between RET and FRS2 is mediated by binding of the phosphotyrosine-binding domain of FRS2 topY1062, a residue in RET that also functions as a binding site for Shc. However, overexpression of FRS2 but not Shc potentiates mitogen-activated protein (MAP) kinase activation by RET oncoproteins. We demonstrate that oncogenic RET-PTC proteins are associated with FRS2 constitutively, leading to tyrosine phosphorylation of FRS2, MAP kinase stimulation, and cell proliferation. However, loss-of-function HSCR-associated RET mutants exhibit impaired FRS2 binding and reduced MAP kinase activation. These experiments demonstrate that FRS2 couples both ligand-regulated and oncogenic forms of RET, with the MAP kinase signaling cascade as part of the response of RET under normal biological conditions and pathological conditions, such as MEN 2 and papillary thyroid carcinomas.

ACKNOWLEDGMENTS

We thank G. Viglietto for the cytofluorimetric analyses and C. Monaco for the RET-PTC molecular constructs. We are grateful to J. Gilder for editing the text.

This study was supported by the Associazione Italiana per la Ricerca sul Cancro (AIRC), by E. C. grant BMH4-CT96–0814, by Programma Biotecnologie legge 95/95 (MURST 5%), and by the Ligue Nationale contre le Cancer grant to M.B. M.S. was supported by a fellowship from FIRC (Italian Foundation for Cancer Research).