ShcA and Grb2 mediate polyoma middle T antigen-induced endothelial transformation and Gab1 tyrosine phosphorylation.
Middle T antigen (PymT) is the principal transforming component of polyomavirus, and rapidly induces hemangiomas in neonatal mice. PymT, a membrane-associated scaffold, recruits and activates Src family tyrosine kinases, and, once tyrosine phosphorylated, binds proteins with PTB and SH2 domains such as ShcA, phosphatidylinositol 3-kinase (PI3K) and phospholipase Cgamma-1 (PLCgamma-1). To explore the pathways required for endothelial transformation in vivo, we introduced PymT mutant forms into mice. PymT variants unable to bind PI3K and PLCgamma-1 directly induced hemangiomas similarly to wild type, but a mutant unable to bind ShcA was transformation compromised. Requirement for a ShcA PTB domain- binding site was suppressed by replacing this motif in PymT with YXN sequences, which bind the Grb2 SH2 domain upon phosphorylation. Surprisingly, PymT recruitment of ShcA and Grb2 correlated with PI3K activation. PymT mimics activated receptor tyrosine kinases by forming a ShcA-Grb2-Gab1 complex, thus inducing Gab1 tyrosine phosphorylation, which itself is associated with PI3K. Therefore, PymT activation of ShcA-Grb2 signaling is critical for endothelial transformation, and PymT can stimulate Grb2 signaling to both the MAP kinase and PI3K pathways.
Ong
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Programme in Molecular Biology and Cancer, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, Ontario M5G 1X5, Canada.
Dilworth
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Hauck-Schmalenberger
I
I
Pawson
T
T
Kiefer
F
F
D016475:3T3 Cells; D048868:Adaptor Proteins, Signal Transducing; D020816:Amino Acid Motifs; D000818:Animals; D000831:Animals, Newborn; D000952:Antigens, Polyomavirus Transforming / Q000378:metabolism*; D001665:Binding Sites; D000081247:CSK Tyrosine-Protein Kinase; D002460:Cell Line; D004247:DNA / Q000378:metabolism; D004727:Endothelium / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D051380:GRB2 Adaptor Protein; D006801:Humans; D020935:MAP Kinase Signaling System; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D010766:Phosphorylation; D011233:Precipitin Tests; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D011916:Rats, Inbred F344; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism; D012190:Retroviridae / Q000235:genetics; D020837:SOS1 Protein / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013997:Time Factors; D014162:Transfection; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases
D048868:Adaptor Proteins, Signal Transducing; D000952:Antigens, Polyomavirus Transforming; D000081247:CSK Tyrosine-Protein Kinase; D004247:DNA; D051380:GRB2 Adaptor Protein; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins; D011505:Protein-Tyrosine Kinases; D011506:Proteins; D020794:Receptor Protein-Tyrosine Kinases; D020837:SOS1 Protein; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D019061:src-Family Kinases; C098587:GAB1 protein, human; C495948:GRB2 protein, human; C490897:Gab1 protein, mouse; C490896:Gab1 protein, rat; C495949:Grb2 protein, mouse; C495950:Grb2 protein, rat; C528024:SHC1 protein, human; C528034:Shc1 protein, mouse; C528035:Shc1 protein, rat; C000631346:CSK protein, human