ERK negatively regulates the epidermal growth factor-mediated interaction of Gab1 and the phosphatidylinositol 3-kinase.
We have examined the ability of epidermal growth factor (EGF)-stimulated ERK activation to regulate Grb2-associated binder-1 (Gab1)/phosphatidylinositol 3-kinase (PI3K) interactions. Inhibiting ERK activation with the MEK inhibitor U0126 increased the EGF-stimulated association of Gab1 with either full-length glutathione S-transferase-p85 or the p85 C-terminal Src homology 2 (SH2) domain, a result reproduced by co-immunoprecipitation of the native proteins from intact cells. This increased association of Gab1 and the PI3K correlates with an increase in PI3K activity and greater phosphorylation of Akt. This result is in direct contrast to what we have previously reported following HGF stimulation where MEK inhibition decreased the HGF-stimulated association of Gab1 and p85. In support of this divergent effect of ERK on Gab1/PI3K association following HGF and EGF stimulation, U0126 decreased the HGF-stimulated association of p85 and the Gab1 c-Met binding domain but did not alter the EGF-stimulated association of p85 and the c-Met binding domain. An examination of the mechanism of this effect revealed that the treatment of cells with EGF + U0126 increased the tyrosine phosphorylation of Gab1 as well as its association with another SH2-containing protein, SHP2. Furthermore, overexpression of a catalytically inactive form of SHP2 or pretreatment with pervanadate markedly increased EGF-stimulated Gab1 tyrosine phosphorylation. These experiments demonstrate that EGF and HGF-mediated ERK activation result in divergent effects on Gab1/PI3K signaling. HGF-stimulated ERK activation increases the Gab1/PI3K association, whereas EGF-stimulated ERK activation results in a decrease in the tyrosine phosphorylation of Gab1 and a decreased association with the PI3K. SHP2 is shown to associate with and dephosphorylate Gab1, suggesting that EGF-stimulated ERK might act through the regulation of SHP2.
Yu
Cheng Fang
CF
Department of Internal Medicine, Section of Nephrology, Yale University School of Medicine, New Haven, Connecticut 06520, USA. chengfang.yu@yale.edu
Liu
Zhen-Xiang
ZX
Cantley
Lloyd G
LG
D048868:Adaptor Proteins, Signal Transducing; D015153:Blotting, Western; D002070:Butadienes / Q000494:pharmacology; D002460:Cell Line; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D015801:Helminth Proteins / Q000378:metabolism; D006801:Humans; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D019859:Proto-Oncogene Proteins c-met / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D015398:Signal Transduction; D014443:Tyrosine / Q000378:metabolism; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D002070:Butadienes; D016895:Culture Media, Serum-Free; D004791:Enzyme Inhibitors; C098587:GAB1 protein, human; D015801:Helminth Proteins; D009570:Nitriles; D010750:Phosphoproteins; D011993:Recombinant Fusion Proteins; C113580:U 0126; C095048:microfilarial sheath protein, Helminth; D014443:Tyrosine; D004815:Epidermal Growth Factor; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D019859:Proto-Oncogene Proteins c-met; D020928:Mitogen-Activated Protein Kinases