Identification of Tek/Tie2 binding partners. Binding to a multifunctional docking site mediates cell survival and migration.
The Tek/Tie2 receptor tyrosine kinase plays a pivotal role in vascular and hematopoietic development. To study the signal transduction pathways that are mediated by this receptor, we have used the yeast two-hybrid system to identify signaling molecules that associate with the phosphorylated Tek receptor. Using this approach, we demonstrate that five molecules, Grb2, Grb7, Grb14, Shp2, and the p85 subunit of phosphatidylinositol 3-kinase can interact with Tek in a phosphotyrosine-dependent manner through their SH2 domains. Mapping of the binding sites of these molecules on Tek reveals the presence of a multisubstrate docking site in the carboxyl tail of Tek (Tyr(1100)). Mutation of this site abrogates binding of Grb2 and Grb7 to Tek in vivo, and this site is required for tyrosine phosphorylation of Grb7 and p85 in vivo. Furthermore, stimulation of Tek-expressing cells with Angiopoietin-1 results in phosphorylation of both Tek and p85 and in activation of endothelial cell migration and survival pathways that are dependent in part on phosphatidylinositol 3-kinase. Taken together, these results demonstrate that Angiopoietin-1-induced signaling from the Tek receptor is mediated by a multifunctional docking site that is responsible for activation of both cell migration and cell survival pathways.
Jones
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N
Division of Cancer Biology Research, Sunnybrook and Women's College Health Sciences Centre, University of Toronto, Toronto, Ontario M4N 3M5.
Master
Z
Z
Jones
J
J
Bouchard
D
D
Gunji
Y
Y
Sasaki
H
H
Daly
R
R
Alitalo
K
K
Dumont
D J
DJ
D048868:Adaptor Proteins, Signal Transducing; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002465:Cell Movement / Q000502:physiology; D002470:Cell Survival / Q000502:physiology; D004622:Embryo, Mammalian; D066246:ErbB Receptors / Q000378:metabolism*; D051380:GRB2 Adaptor Protein; D051382:GRB7 Adaptor Protein; D015723:Gene Library; D047908:Intracellular Signaling Peptides and Proteins; D008168:Lung / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009206:Myocardium / Q000378:metabolism; D019869:Phosphatidylinositol 3-Kinases / Q000737:chemistry / Q000378:metabolism; D011485:Protein Binding; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D042787:Receptor, TIE-2; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development; D018909:src Homology Domains
D048868:Adaptor Proteins, Signal Transducing; D066246:ErbB Receptors; D051380:GRB2 Adaptor Protein; D051382:GRB7 Adaptor Protein; D047908:Intracellular Signaling Peptides and Proteins; D019869:Phosphatidylinositol 3-Kinases; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D053824:Protein Tyrosine Phosphatase, Non-Receptor Type 6; D017027:Protein Tyrosine Phosphatases; D011506:Proteins; D020794:Receptor Protein-Tyrosine Kinases; D042787:Receptor, TIE-2; D011993:Recombinant Fusion Proteins; D054593:SH2 Domain-Containing Protein Tyrosine Phosphatases; C490977:Grb14 protein, mouse; C495949:Grb2 protein, mouse; C495956:Grb7 protein, mouse; C516725:Ptpn11 protein, mouse; C509248:Ptpn6 protein, mouse