Biochemical and biological responses induced by coupling of Gab1 to phosphatidylinositol 3-kinase in RET-expressing cells.
Grb2-associated binder-1 (Gab1) is a docking protein closely related to insulin receptor substrates. We previously reported that tyrosine 1062 in RET receptor tyrosine kinase activated by glial cell line-derived neurotrophic factor (GDNF) represents a binding site for the Shc-Grb2-Gab1 complex, and that the p85 subunit of phosphatidylinositol 3-kinase (PI3K) and SHP2 tyrosine phosphatase is associated with Gab1 in GDNF-treated cells. In the present study, we further analyzed the physiological roles of Gab1 downstream of RET, using Gab1 mutants that lack the binding sites for PI3K (Gab1 PI3K-m) or SHP-2 (Gab1 SHP2-m). Expression of Gab1 PI3K-m in SK-N-MC human primitive neuroectodermal tumor cells expressing wild-type RET markedly impaired Akt phosphorylation, Rac1 activation, and lamellipodia formation that were induced by GDNF whereas expression of Gab1 SHP2-m partially impaired Erk activation. Furthermore, expression of Gab1 PI3K-m, but not Gab1 SHP2-m, in TT human medullary thyroid carcinoma cells expressing RET with a multiple endocrine neoplasia 2A mutation enhanced cytochrome c release, and apoptosis induced by etoposide, suggesting that PI3K is involved in survival of TT cells via a mitochondrial pathway. These findings demonstrated that coupling of Gab1 to PI3K is important for biological responses in RET-expressing cells.
Maeda
Kengo
K
Department of Pathology, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan.
Murakami
Hideki
H
Yoshida
Reiko
R
Ichihara
Masatoshi
M
Abe
Akihiro
A
Hirai
Makoto
M
Murohara
Toyoaki
T
Takahashi
Masahide
M
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D017209:Apoptosis; D001665:Binding Sites; D045744:Cell Line, Tumor; D002469:Cell Separation; D045304:Cytochromes c / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005047:Etoposide / Q000494:pharmacology; D005434:Flow Cytometry; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D005982:Glutathione Transferase / Q000378:metabolism; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D051379:Mice; D008856:Microscopy, Fluorescence; D008928:Mitochondria / Q000378:metabolism; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008954:Models, Biological; D008957:Models, Genetic; D009154:Mutation; D041681:NIH 3T3 Cells; D009414:Nerve Growth Factors / Q000737:chemistry; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010750:Phosphoproteins / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection
D048868:Adaptor Proteins, Signal Transducing; D018076:DNA, Complementary; C098587:GAB1 protein, human; C495039:GDNF protein, human; C495040:Gdnf protein, mouse; D051100:Glial Cell Line-Derived Neurotrophic Factor; D051097:Glial Cell Line-Derived Neurotrophic Factor Receptors; D009414:Nerve Growth Factors; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D005047:Etoposide; D045304:Cytochromes c; D005982:Glutathione Transferase; D019869:Phosphatidylinositol 3-Kinases; D051096:Proto-Oncogene Proteins c-ret; D020794:Receptor Protein-Tyrosine Kinases; C494999:Ret protein, mouse; D020928:Mitogen-Activated Protein Kinases