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Sci. Signal., 26 June 2012
Vol. 5, Issue 230, p. ec177
[DOI: 10.1126/scisignal.2003339]

EDITORS' CHOICE

Cell Biology Grb2: Adapting to Basal Activity

Nancy R. Gough

Science Signaling, AAAS, Washington, DC 20005, USA

Fibroblast growth factor receptors (FGFRs) are receptor tyrosine kinases that initiate a phosphorylation cascade in response to ligand binding. Grb2 is an adaptor protein with N- and C-terminal SH3 domains, which bind to proline-rich regions, separated by an SH2 domain that binds to phosphorylated tyrosines. Grb2 has been considered a positive player in the FGFR pathway through its ability to bind to the phosphorylated and activated receptor through the SH2 domain and recruit other proteins to the receptor for activation through the SH3 domains. Lin et al. now suggest that Grb2 can also function as a dimer and, through the SH3 domains, bind to two unphosphorylated FGFRs. This resulted in receptor dimerization and basal receptor phosphorylation, while preventing downstream signaling. In cells expressing a mutant of FGFR2 that could not be stimulated by ligands, the amount of basal receptor phosphorylation increased with increasing amounts of coexpressed Grb2. In vitro phosphorylation of the cytoplasmic domain of FGFR was enhanced by the presence of increasing amounts of Grb2. Molecular modeling suggested that Grb2 bound to the FGFR through the SH3 domain, and isothermal titration calorimetry (ITC) showed that Grb2 and the FGFR cytoplasmic domain interacted initially as a 2:1 complex and then, as more FGFR was added, with equal stoichiometry. Dynamic light scattering indicated the formation of a 2:2 heterotetramer, and in vitro fluorescence resonance energy transfer (FRET) with labeled FGFR cytoplasmic domains indicated that Grb2 facilitated FGFR dimerization. The molecular modeling placed a phosphorylatable tyrosine on the receptor in close proximity to a phosphorylatable tyrosine on Grb2, and in vitro kinase studies with various truncation mutants of Grb2 showed that this tyrosine in the Grb2 SH3 domain was a substrate for FGFR. Phosphorylation of the receptor and this tyrosine on Grb2 is predicted to cause Grb2 to dissociate from the receptor, and in vitro ITC studies confirmed that the phosphorylated receptor cytoplasmic domain and phosphorylated Grb2 failed to interact. In transfected cells, Grb2 with a phenylalanine substitution at this tyrosine inhibited ligand-activated FGFR signaling without affecting basal phosphorylation, consistent with mutant Grb2 remaining associated with the receptor. Thus, Grb2 may have a dual function in receptor tyrosine kinase signaling—first, as an inhibitory adaptor that primes the receptors by promoting dimerization and receptor phosphorylation, while preventing downstream signaling, and then second, as a positive adaptor, recruiting proteins to the ligand-stimulated receptor to promote downstream signaling.

C.-C. Lin, F. A. Melo, R. Ghosh, K. M. Suen, L. J. Stagg, J. Kirkpatrick, S. T. Arold, Z. Ahmed, J. E. Ladbury, Inhibition of basal FGF receptor signaling by dimeric Grb2. Cell 149, 1514–1524 (2012). [Online Journal]

Citation: N. R. Gough, Grb2: Adapting to Basal Activity. Sci. Signal. 5, ec177 (2012).



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