Dok-3 sequesters Grb2 and inhibits the Ras-Erk pathway downstream of protein-tyrosine kinases.
Adaptor proteins are essential in coordinating recruitment and, in a few cases, restraint of various effectors during cellular signaling. Dok-1, Dok-2 and Dok-3 comprise a closely related family of adaptor, which negatively regulates mitogen-activated protein kinase Erk downstream of protein-tyrosine kinases (PTKs). Recruitment of p120 rasGAP, a potent inhibitor of Ras, by Dok-1 and Dok-2 appears critical in the negative regulation of the Ras-Erk pathway. However, as Dok-3 does not bind rasGAP, it has been unclear how Dok-3 inhibits Erk downstream of PTKs. Here, we identified Grb2 as a Dok-3-binding protein upon its tyrosine phosphorylation. This interaction required the intact binding motifs of the Grb2 SH2 domain, and a mutant (Dok-3-FF) having a Tyr/Phe substitution at these motifs failed to inhibit Ras and Erk activation downstream of a cytoplasmic PTK Src. Because Grb2 forms a stable complex with Sos, a crucial activator of Ras, these data suggest that Dok-3 restrains Grb2 and inhibits the ability of the Grb2-Sos complex to activate Ras. Indeed, forced expression of Dok-3, but not Dok-3-FF, inhibited the recruitment of the Grb2-Sos complex to Shc downstream of Src, which is an essential event for activation of the Ras-Erk pathway. These findings indicate that Dok-3 sequesters Grb2 from Shc and inhibits the Ras-Erk pathway downstream of PTKs.
Honma
Miyuki
M
Department of Cell Regulation, Medical Research Institute, Tokyo Medical and Dental University, Tokyo 113-8510, Japan.
Higuchi
Osamu
O
Shirakata
Masaki
M
Yasuda
Tomoharu
T
Shibuya
Hiroshi
H
Iemura
Shun-ichiro
S
Natsume
Tohru
T
Yamanashi
Yuji
Y
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D048049:Extracellular Signal-Regulated MAP Kinases / Q000037:antagonists & inhibitors* / Q000378:metabolism; D051380:GRB2 Adaptor Protein / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D051379:Mice; D008969:Molecular Sequence Data; D015688:Oncogene Protein pp60(v-src) / Q000378:metabolism; D011485:Protein Binding; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D016283:Proto-Oncogene Proteins p21(ras) / Q000037:antagonists & inhibitors* / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins; D015398:Signal Transduction; D020664:Son of Sevenless Proteins / Q000378:metabolism; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C518789:DOK3 protein, human; D051380:GRB2 Adaptor Protein; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; C528034:Shc1 protein, mouse; D020664:Son of Sevenless Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D014443:Tyrosine; D011505:Protein-Tyrosine Kinases; D015688:Oncogene Protein pp60(v-src); D048049:Extracellular Signal-Regulated MAP Kinases; D016283:Proto-Oncogene Proteins p21(ras)