The Shb adaptor protein binds to tyrosine 766 in the FGFR-1 and regulates the Ras/MEK/MAPK pathway via FRS2 phosphorylation in endothelial cells.
Stimulation of fibroblast growth factor receptor-1 (FGFR-1) is known to result in phosphorylation of tyrosine 766 and the recruitment and subsequent activation of phospholipase C-gamma (PLC-gamma). To assess the role of tyrosine 766 in endothelial cell function, we generated endothelial cells expressing a chimeric receptor, composed of the extracellular domain of the PDGF receptor-alpha and the intracellular domain of FGFR-1. Mutation of tyrosine 766 to phenylalanine prevented PLC-gamma activation and resulted in a reduced phosphorylation of FRS2 and reduced activation of the Ras/MEK/MAPK pathway relative to the wild-type chimeric receptor. However, FGFR-1-mediated MAPK activation was not dependent on PKC activation or intracellular calcium, both downstream mediators of PLC-gamma activation. We report that the adaptor protein Shb is also able to bind tyrosine 766 in the FGFR-1, via its SH2 domain, resulting in its subsequent phosphorylation. Overexpression of an SH2 domain mutant Shb caused a dramatic reduction in FGFR-1-mediated FRS2 phosphorylation with concomitant perturbment of the Ras/MEK/MAPK pathway. Expression of the chimeric receptor mutant and the Shb SH2 domain mutant resulted in a similar reduction in FGFR-1-mediated mitogenicity. We conclude, that Shb binds to tyrosine 766 in the FGFR-1 and regulates FGF-mediated mitogenicity via FRS2 phosphorylation and the subsequent activation of the Ras/MEK/MAPK pathway.
Department of Genetics and Pathology, Uppsala University, Rudbeck Laboratory, Sweden. Michael.Cross@genpat.uu.se
Michael J
MJ
Cross
Lingge
L
Lu
Peetra
P
Magnusson
Daniel
D
Nyqvist
Kristina
K
Holmqvist
Michael
M
Welsh
Lena
L
Claesson-Welsh
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D002460:Cell Line; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism; D015801:Helminth Proteins / Q000378:metabolism; D048728:MAP Kinase Kinase Kinase 1; D020935:MAP Kinase Signaling System / Q000502:physiology*; D008565:Membrane Proteins / Q000378:metabolism*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation; D051966:Phospholipase C gamma; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011485:Protein Binding; D011493:Protein Kinase C / Q000378:metabolism; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D020794:Receptor Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D051496:Receptor, Fibroblast Growth Factor, Type 1; D020796:Receptor, Platelet-Derived Growth Factor alpha / Q000235:genetics / Q000378:metabolism; D017468:Receptors, Fibroblast Growth Factor / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D010738:Type C Phospholipases / Q000378:metabolism; D014443:Tyrosine / Q000378:metabolism*; D018631:ras Proteins / Q000378:metabolism; D018909:src Homology Domains; D019061:src-Family Kinases / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C106534:FRS2 protein, human; D015801:Helminth Proteins; D008565:Membrane Proteins; D010750:Phosphoproteins; D010982:Platelet-Derived Growth Factor; D011518:Proto-Oncogene Proteins; D017468:Receptors, Fibroblast Growth Factor; D011993:Recombinant Fusion Proteins; C490081:SHB protein, human; C085634:Shb protein, mouse; C075278:platelet-derived growth factor A; D016222:Fibroblast Growth Factor 2; C095048:microfilarial sheath protein, Helminth; D014443:Tyrosine; C496349:Fgfr1 protein, mouse; D020794:Receptor Protein-Tyrosine Kinases; D051496:Receptor, Fibroblast Growth Factor, Type 1; D020796:Receptor, Platelet-Derived Growth Factor alpha; D019061:src-Family Kinases; D017346:Protein Serine-Threonine Kinases; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D048728:MAP Kinase Kinase Kinase 1; C482576:MAP3K1 protein, human; C482578:Map3k1 protein, mouse; D010738:Type C Phospholipases; D051966:Phospholipase C gamma; D018631:ras Proteins