Point mutation in FGF receptor eliminates phosphatidylinositol hydrolysis without affecting mitogenesis

Abstract

Stimulation of growth factor receptors with tyrosine kinase activity is followed by rapid receptor dimerization, tyrosine autophosphorylation and phosphorylation of signalling molecules such as phospholipase C gamma (PLC gamma) and the ras GTPase-activating protein. PLC gamma and GTPase-activating protein bind to specific tyrosine-phosphorylated regions in growth factor receptors through their src-homologous SH2 domains. Growth factor-induced tyrosine phosphorylation of PLC gamma is essential for stimulation of phosphatidylinositol hydrolysis in vitro and in vivo. We have shown that a short phosphorylated peptide containing tyrosine at position 766 from a conserved region of the fibroblast growth factor (FGF) receptor is a binding site for the SH2 domain of PLC gamma (ref. 8). Here we show that an FGF receptor point mutant in which Tyr 766 is replaced by a phenylalanine residue (Y766F) is unable to associate with and tyrosine-phosphorylate PLC gamma or to stimulate hydrolysis of phosphatidylinositol. Nevertheless, the Y766F FGF receptor mutant can be autophosphorylated, and can phosphorylate several cellular proteins and stimulate DNA synthesis. Our data show that phosphorylation of the conserved Tyr 766 of the FGF receptor is essential for phosphorylation of PLC gamma and for hydrolysis of phosphatidylinositol, but that elimination of this hydrolysis does not affect FGF-induced mitogenesis.

Cell growth control by G protein-coupled receptors: from signal transduction to signal integration

“…By contrast, many studies have implicated phospholipase C-b activation and enhanced phosphatidylinositol bisphosphate (PIP 2 ) hydrolysis in mitogenesis (Rozengurt, 1986). However, experiments utilizing mutant tyrosine kinase receptors provided evidence that PIP 2 hydrolysis may be neither necessary nor su cient for mitogenesis (Coughlin et al, 1989;Mohammadi et al, 1992). Consistent with those observations, a number of agonists acting on GPCRs, can e ectively induce PIP 2 -hydrolysis, but fail to stimulate growth when added alone to quiescent cells (Moolenaar, 1991).…”

Contribution of Src and Ras pathways in FGF-2 induced endothelial cell differentiation

“…Phosphorylated tyrosine residues, and surrounding amino acids, serve as binding sites for proteins with Src homology (SH)2 domains. In the FGFR-1, only Tyr766 have been identi®ed as a direct binding site for an SH2 containing protein; phospholipase C-g (PLC-g) (Mohammadi et al, 1992;Peters et al, 1992). The role of PLC-g in FGFR-1 signaling is not clear, however, inhibition of PLC-g activation by mutation of the binding site in the FGFR-1 partly reduced the FGF-2 induced mitogenicity (Klint et al, 1995), and PLC-g binding and activation by FGFR-1 has been implicated in regulation of Src kinase activity (Landgren et al, 1995).…”

Fibroblast growth factor receptor-1 mediates chemotaxis independently of direct SH2-domain protein binding

“…The biological consequences of this activation are not completely understood. The Y766F mutant FGFR-1 in which tyrosine 766 is replaced by a phenylalanine residue, internalizes at a reduced rate (Sorokin et al, 1994) and causes a reduced MAP kinase activation (Huang et al, 1995), but is both mitogenically and chemotactically competent (Clyman et al, 1994;Mohammadi et al, 1992;Peters et al, 1992). Activation of PLC-g is also not critical for induction of urokinase type plasminogen activator (uPa; Roghani et al, 1996).…”