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A major Grb2-associated binder-1 (Gab1) binding partner in epidermal growth factor (EGF)-stimulated cells is protein-tyrosine phosphatase (PTPase) SHP2, which contains tandem SH2 domains. The SHP2 PTPase activity is required for activation of the extracellular signal-regulated kinase (ERK) subfamily of mitogen-activated protein (MAP) kinase by EGF. To investigate the mechanism by which Gab1 and SHP2 mediate ERK activation, we characterized the Gab1-SHP2 interaction. We found that both Tyr-627 and Tyr-659 of Gab1 were required for SHP2 binding to Gab1 and for ERK2 activation by EGF. Far Western blot analysis suggested that the tandem SH2 domains of SHP2 bind to Gab1 in a specific orientation, in which the N-SH2 domain binds to phosphotyrosine (Tyr(P))-627 and the C-SH2 domain binds to Tyr(P)-659. When assayed with peptide substrates, SHP2 PTPase was activated by a bisphosphopeptide containing both Tyr(P)-627 and Tyr(P)-659, but not by monophosphopeptides containing Tyr(P)-627 or Tyr(P)-659 or a mixture of these monophosphopeptides. These results suggest that Tyr(P)-627 and Tyr(P)-659 of Gab1 constitute a bisphosphoryl tyrosine-based activation motif (BTAM) that binds and activates SHP2. Remarkably, while a constitutively active SHP2 (SHP2ΔN) could not rescue the defect of a SHP2-binding defective Gab1 (Gab1FF) in ERK2 activation, expression of a Gab1FF-SHP2ΔN chimera resulted in constitutive activation of ERK2 in transfected cells. Thus, physical association of activated SHP2 with Gab1 is necessary and sufficient to mediate the ERK mitogen-activated protein kinase activation. Phosphopeptides derived from Gab1 were dephosphorylated by active SHP2 in vitro. Consistently, substrate-trapping experiments with a SHP2 catalytic inactive mutant suggested that Gab1 was a SHP2 PTPase substrate in the cells. Therefore, Gab1 not only is a SHP2 activator but also is a target of its PTPase. A major Grb2-associated binder-1 (Gab1) binding partner in epidermal growth factor (EGF)-stimulated cells is protein-tyrosine phosphatase (PTPase) SHP2, which contains tandem SH2 domains. The SHP2 PTPase activity is required for activation of the extracellular signal-regulated kinase (ERK) subfamily of mitogen-activated protein (MAP) kinase by EGF. To investigate the mechanism by which Gab1 and SHP2 mediate ERK activation, we characterized the Gab1-SHP2 interaction. We found that both Tyr-627 and Tyr-659 of Gab1 were required for SHP2 binding to Gab1 and for ERK2 activation by EGF. Far Western blot analysis suggested that the tandem SH2 domains of SHP2 bind to Gab1 in a specific orientation, in which the N-SH2 domain binds to phosphotyrosine (Tyr(P))-627 and the C-SH2 domain binds to Tyr(P)-659. When assayed with peptide substrates, SHP2 PTPase was activated by a bisphosphopeptide containing both Tyr(P)-627 and Tyr(P)-659, but not by monophosphopeptides containing Tyr(P)-627 or Tyr(P)-659 or a mixture of these monophosphopeptides. These results suggest that Tyr(P)-627 and Tyr(P)-659 of Gab1 constitute a bisphosphoryl tyrosine-based activation motif (BTAM) that binds and activates SHP2. Remarkably, while a constitutively active SHP2 (SHP2ΔN) could not rescue the defect of a SHP2-binding defective Gab1 (Gab1FF) in ERK2 activation, expression of a Gab1FF-SHP2ΔN chimera resulted in constitutive activation of ERK2 in transfected cells. Thus, physical association of activated SHP2 with Gab1 is necessary and sufficient to mediate the ERK mitogen-activated protein kinase activation. Phosphopeptides derived from Gab1 were dephosphorylated by active SHP2 in vitro. Consistently, substrate-trapping experiments with a SHP2 catalytic inactive mutant suggested that Gab1 was a SHP2 PTPase substrate in the cells. Therefore, Gab1 not only is a SHP2 activator but also is a target of its PTPase. protein-tyrosine phosphatase Grb2-associated binder-1 tyrosine-based activation motif bisphosphoryl TAM Src homology 2 phosphotyrosine mitogen-activated protein kinase extracellular signal-regulated kinase 2 (p42MAPK) epidermal growth factor myelin basic protein NH2-terminal Src homology 2 domain COOH-terminal Src homology 2 domain glutathioneS-transferase hemagglutinin. IRS-1, insulin receptor substrate-1 SHP2 is a protein-tyrosine phosphatase (PTPase)1 with two Src homology-2 (SH2) domains (1Feng G-S. Exp. Cell Res. 1999; 253: 47-54Crossref PubMed Scopus (254) Google Scholar, 2Neel B.G. Tonks N.K. Curr. Opin. Cell Biol. 1998; 9: 193-204Crossref Scopus (742) Google Scholar). These two SH2 domains, termed N-SH2 and C-SH2 domains, are arranged in tandem at the amino (N)-terminal portion of the protein. SHP2 has a low basal PTPase activity that can be activated by deletion of N-SH2 or both SH2 domains or by specific phosphopeptides that bind to the SH2 domains. The crystal structure of SHP2 shows that the N-SH2 domain is inserted into the catalytic cleft of the phosphatase domain in the absence of a ligand for the N-SH2 domain, thus maintaining the phosphatase in an autoinhibitory state (3Hof P. Pluskey S. Dhe-Paganon S. Eck M.J. Shoelson S.E. Cell. 1998; 92: 441-450Abstract Full Text Full Text PDF PubMed Scopus (775) Google Scholar). Binding of a specific tyrosine-based activation motif (TAM) to the N-SH2 domain is predicted to induce an allosteric change that disrupts the inhibitory interaction between the N-SH2 and the catalytic domains, leading to phosphatase activation (3Hof P. Pluskey S. Dhe-Paganon S. Eck M.J. Shoelson S.E. Cell. 1998; 92: 441-450Abstract Full Text Full Text PDF PubMed Scopus (775) Google Scholar). Although there is no direct surface contact between the C-SH2 and the catalytic domains, the C-SH2 domain connects the N-SH2 domain to the PTPase domain and contributes to the selectivity and high affinity binding of the tandem SH2 domains to bisphosphoryl TAMs (4Eck M.J. Pluskey S. Trub T. Harrison S.C. Shoelson S.E. Nature. 1996; 379: 277-280Crossref PubMed Scopus (171) Google Scholar, 5Ottinger E.A. Botfield M.C. Shoelson S.E. J. Biol. Chem. 1998; 273: 729-735Abstract Full Text Full Text PDF PubMed Scopus (184) Google Scholar, 6Ohnishi H. Kubota M. Ohtake A. Sato K. Sano S. J. Biol. Chem. 1996; 271: 25569-25574Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar). It has been demonstrated that the SHP2 PTPase activity is required for activation of the extracellular signal-regulated protein kinase (ERK) subfamily of mitogen-activated protein (MAP) kinase by epidermal growth factor (EGF) (7Deb T.B. Wong L. Salomon D.S. Zhou G. Dixon J.E. Gutkind S. Thompson S.A. Johnson G.R. J. Biol. Chem. 1998; 273: 16643-16646Abstract Full Text Full Text PDF PubMed Scopus (82) Google Scholar, 8Bennett A.M. Hausdorff S.F. O'Reilly A.M. Freeman R.M. Neel B.G. Mol. Cell. Biol. 1996; 16: 1189-1202Crossref PubMed Scopus (226) Google Scholar). However, the mechanism by which SHP2 exerts its positive role in ERK MAP kinase activation by EGF is not yet known. The autoinhibitory configuration of the unbound SHP2 necessitates a binding partner for activation of the SHP2 PTPase. Although insulin receptor substrate-1 (IRS-1) and SHP substrate-1 (SHPS-1, also called BIT, SIRPα) can bind and activate SHP2 (5Ottinger E.A. Botfield M.C. Shoelson S.E. J. Biol. Chem. 1998; 273: 729-735Abstract Full Text Full Text PDF PubMed Scopus (184) Google Scholar, 6Ohnishi H. Kubota M. Ohtake A. Sato K. Sano S. J. Biol. Chem. 1996; 271: 25569-25574Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar), IRS-1 is not tyrosine phosphorylated in response to EGF (9Kadowaki T. Koyasu S. Nishida E. Tobe K. Izumi T. Takaku F. Sakai H. Yahara I. Kasuga M. J. Biol. Chem. 1987; 262: 7342-7344Abstract Full Text PDF PubMed Google Scholar). Moreover, the interaction between IRS-1 and SHP2 is not required for ERK activation by insulin (10Myers M.G. Mendez R. Shi P. Pierce J.H. Rhoadst R. White M. J. Biol. Chem. 1998; 273: 26908-26914Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar). In fact, interaction between IRS-1 and SHP2 appears to have negative effects on insulin signaling (10Myers M.G. Mendez R. Shi P. Pierce J.H. Rhoadst R. White M. J. Biol. Chem. 1998; 273: 26908-26914Abstract Full Text Full Text PDF PubMed Scopus (140) Google Scholar). Similarly, it has been reported that SHPS-1 and its interaction with SHP2 play a negative role in the EGF-induced ERK activation pathway (11Kharitonenkov A. Chen Z. Sures I. Wang H. Chilling J. Ullrich A. Nature. 1997; 386: 181-186Crossref PubMed Scopus (563) Google Scholar). A major SHP2 binding partner in EGF-stimulated cells is Grb2-associated binder-1 (Gab1) (12Holgado-Madruga M. Emlet D.R. Moscatello D.K. Godwin A.K. Wong A.J. Nature. 1996; 379: 560-564Crossref PubMed Scopus (605) Google Scholar). Gab1 is a multisite docking protein containing an N-terminal pleckstrin homology (PH) domain and several proline-rich SH3 domain-binding sequences (12Holgado-Madruga M. Emlet D.R. Moscatello D.K. Godwin A.K. Wong A.J. Nature. 1996; 379: 560-564Crossref PubMed Scopus (605) Google Scholar, 13Weidner K.M. Cesare S.D. Sachs M. Brinkmann V. Behrens J. Birchmeier W. Nature. 1996; 384: 173-176Crossref PubMed Scopus (509) Google Scholar). Upon stimulation of cells with EGF and other growth factors and cytokines, Gab1 becomes phosphorylated on multiple tyrosine residues, which bind SHP2, and other SH2 (12Holgado-Madruga M. Emlet D.R. Moscatello D.K. Godwin A.K. Wong A.J. Nature. 1996; 379: 560-564Crossref PubMed Scopus (605) Google Scholar, 13Weidner K.M. Cesare S.D. Sachs M. Brinkmann V. Behrens J. Birchmeier W. Nature. 1996; 384: 173-176Crossref PubMed Scopus (509) Google Scholar, M. T. T. E. K. M. T. Mol. Cell. Biol. 1998; PubMed Scopus Google Scholar, T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, M. Z. J. Mol. Cell. Biol. PubMed Scopus Google Scholar, M. M. G-S. Mol. Cell. Biol. PubMed Scopus Google Scholar, S. S. J. M. Wong A.J. E. 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These to Gab1 a SHP2 binding partner that EGF-stimulated ERK activation. We found that Gab1 with a at Tyr-627 SHP2 binding activity and EGF-stimulated ERK2 (p42MAPK) activation T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). However, the mechanism by which the Gab1-SHP2 interaction ERK activation has not been In the we that two tyrosine and in the of Gab1 are both required for SHP2 binding to Gab1 and for EGF-stimulated ERK activation. These two tyrosine constitute motif (BTAM) that mediate binding and activation of SHP2 by of a constitutive active SHP2 (SHP2ΔN) not rescue the defect of a SHP2 Gab1 expression of a Gab1FF-SHP2ΔN protein resulted in constitutive activation of ERK2 in cells. These that the activated SHP2 has to be with Gab1 to mediate EGF-stimulated ERK2 activation and Gab1 the SHP2 activator for the ERK MAP kinase pathway in EGF-stimulated cells. in experiments and substrate-trapping in cells suggest that Gab1 is a substrate of the SHP2 PTPase. were from the from from from from and and from basic protein and were from was from of SHP2 N-SH2 domain and C-SH2 domain have been M. G-S. L. J. 1999; PubMed Google Scholar). of the and N-SH2 domain deletion of SHP2 were reported M. G-S. L. J. 1999; PubMed Google Scholar, L. J. Biol. Chem. Full Text PDF PubMed Google Scholar). The expression for the SHP2, a SHP2 the N-SH2 domain and a catalytic inactive to mutant of SHP2 have also been reported M. G-S. L. J. 1999; PubMed Google Scholar). and were by I. of a derived from the was from and were and by The amino sequences of these are phosphotyrosine The of other were T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google or for Gab1 and have been T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). for Gab1 with a Tyr-659 to and Gab1 with of both Tyr-627 and Tyr-659 to (Gab1FF) were with the in and The was are were by To a for the expression of a Gab1FF-SHP2ΔN a for amino of SHP2 L. J. Biol. Chem. Full Text PDF PubMed Google was by The for a and The was inserted into a between the of and the in cells were in with was on cells in to the cells were with in for The were from the and the cells were with growth the cells were in with for and for were in A 2 2 and and Gab1 were with the were on to and for analysis T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). Far Western blot analysis of binding to the and of these Gab1 were on and to The were with in for and with in the for The were and with an and a and with the substrate was by the with an with and of the with the were in and in A containing and was with a the of was to ERK2 while the other was for ERK2 protein by the kinase the were for at in of kinase myelin basic protein containing of The was by the of and at for The were on The were by the were with a The ERK2 activity of was by the of with that of the or which been A ERK2 was for analysis of ERK2 protein in the The of were by and by the To the of was in of and at for in a H. M. S. PubMed Scopus Google Scholar, P. Wong R. I. J. PubMed Scopus Google Scholar). of was and of mixture was to the of by the A of was that been the the phosphopeptides P. Wong R. I. J. PubMed Scopus Google Scholar). SHP2 PTPase activity was by the from phosphopeptides the P. Wong R. I. J. PubMed Scopus Google Scholar). The PTPase was at for in a mixture containing and the of phosphopeptides and PTPase. The were that of the substrate was dephosphorylated by the of the The was by of of P. Wong R. I. J. PubMed Scopus Google Scholar). for at the of the mixture at was in a with a A of was in to the PTPase Although the was between and of a was in the We that Gab1 with a Tyr-627 to SHP2 binding activity in EGF-stimulated that Tyr-627 of Gab1 is in Gab1-SHP2 interaction T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). of the Gab1 amino that Tyr-659 of Gab1 a to be phosphorylated by the EGF receptor S. J. A. E. V. W. 1999; PubMed Scopus Google Scholar), is also in a binding for the SH2 domain of SHP2. To tyrosine with Tyr-627 to with the tandem SH2 domains of SHP2, we the effects of and of Tyr-659 and Tyr-627 on SHP2 binding to or Gab1 was in cells by Gab1-SHP2 binding was in cells with or EGF by analysis of Gab1 with an SHP2 was with Gab1 in cells in which Gab1 was not Upon EGF tyrosine in Gab1 phosphorylated and SHP2 was to the Gab1 to of Gab1 at Tyr-627 or Tyr-659 or both (Gab1FF) resulted in of the SHP2 binding activity A that of Gab1 were in Gab1 that both Tyr-627 and Tyr-659 are required for SHP2 binding to Gab1 in response to EGF. analysis with an that and of phosphotyrosine that of the Gab1 in EGF-stimulated cells that Tyr-627 and Tyr-659 were phosphorylated in the Gab1 in EGF-stimulated in with a S. J. A. E. V. W. 1999; PubMed Scopus Google Scholar). of was in EGF-stimulated with the that tyrosine Tyr-627 and Tyr-659 were phosphorylated EGF stimulation T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, S. J. A. E. V. W. 1999; PubMed Scopus Google Scholar). To Tyr-659 is required for EGF-stimulated ERK2 activation, and were with ERK2 in and EGF-stimulated ERK2 activation was in EGF resulted in a activation of of the Gab1 on EGF-stimulated ERK2 activation, while expression of a inhibitory on EGF-stimulated ERK2 activation reported T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). to expression of or EGF-stimulated ERK2 activation. Thus, both Tyr-627 and Tyr-659 of Gab1 are necessary for Gab1 to mediate the EGF-stimulated ERK2 activation. To the interaction between Gab1 and SHP2, we Far Western blot analysis containing the N-SH2 or C-SH2 domain of SHP2. and were from EGF-stimulated cells or The were on to and with or by an the were with an or an The binding of protein to Gab1 was In and to the Gab1 in an but not to and to with a but not to that the N-SH2 domain of SHP2 binds to the Tyr(P)-627 in Gab1 A and the other to A and These results on in that both SH2 domains of SHP2 could with Gab1 in EGF-stimulated cells. These also suggest that SHP2 with Gab1 in a specific in which the N-SH2 domain binds the Tyr(P)-627 while the C-SH2 domain binds the Tyr(P)-659 To phosphopeptides containing Tyr-627 Tyr-659 can activate the PTPase activity of SHP2, we assayed the SHP2 PTPase activity two phosphopeptides The was a SHP2 substrate in contains the autoinhibitory of have peptide a substrate of constitutively active SHP2 that not SHP2 activity H. Kubota M. Ohtake A. Sato K. Sano S. J. Biol. Chem. 1996; 271: 25569-25574Abstract Full Text Full Text PDF PubMed Scopus (86) Google Scholar, M. I. F. J. Biol. Chem. Full Text PDF PubMed Google Scholar). in A the the was dephosphorylated at a of by SHP2 of monophosphopeptides containing Tyr(P)-627 or Tyr(P)-659 resulted in a in the change in was to of or by SHP2, the was the of the of and or were with SHP2. Therefore, there was only an in the that and not activate the SHP2 PTPase. results were with of and that we have to or not Similarly, a mixture of and could not activate SHP2 at that we have and not We the of a bisphosphopeptide on SHP2 is a bisphosphoryl peptide containing Tyr(P)-627 and Tyr(P)-659 of Gab1 and the amino between and Tyr(P)-627 and Tyr(P)-659. in of to the mixture resulted in activation of SHP2. of SHP2 by was The activity of of Tyr-627 and a peptide with amino not have on SHP2 A and and not In fact, the peptide was a negative with in experiments in these that can activate SHP2 PTPase and that of on both of Tyr-627 and Tyr-659 and the physical of these two phosphotyrosine in a To that can activate SHP2, we the SHP2 PTPase activity a contains which is of the in Gab1 T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, S. S. J. M. Wong A.J. E. Mol. 1998; PubMed Scopus Google and a target of SHP2 in the cells. in we found that SHP2 was not activated by monophosphopeptides a mixture of and or the peptide of the bisphosphopeptide to the mixture resulted in activation of SHP2 PTPase To the that the activation of SHP2 by was to of the phosphatase and we the of these phosphopeptides by a constitutively active SHP2. N-SH2 domain deletion mutant of SHP2 (SHP2ΔN) was for results were in at and peptide in the active SHP2 dephosphorylated but at a the of SHP2 or was the could not be to a of the active SHP2 dephosphorylated yet not the SHP2 PTPase activity the that activation of the SHP2 PTPase activity by in A was to of in that a of Gab1-SHP2 interaction is activation of SHP2 PTPase. SHP2 PTPase activity is required for ERK2 activation in response to EGF stimulation (7Deb T.B. Wong L. Salomon D.S. Zhou G. Dixon J.E. Gutkind S. Thompson S.A. Johnson G.R. J. Biol. Chem. 1998; 273: 16643-16646Abstract Full Text Full Text PDF PubMed Scopus (82) Google Scholar, 8Bennett A.M. Hausdorff S.F. O'Reilly A.M. Freeman R.M. Neel B.G. Mol. Cell. Biol. 1996; 16: 1189-1202Crossref PubMed Scopus (226) Google Scholar), we activation of SHP2 is the only of Gab1-SHP2 interaction. To we expression of a constitutively active SHP2 could rescue the inhibitory of which binds SHP2 is predicted to activate its PTPase in the N-SH2 domain deletion mutant of SHP2 (SHP2ΔN) a constitutively activated PTPase which could not be activated by the Gab1 bisphosphoryl peptide The SHP2 or was with the Gab1 or in cells and EGF-stimulated ERK2 activation in the transfected cells was of SHP2 or with the Gab1 on EGF-stimulated ERK2 activation 2 and in has a inhibitory on EGF-stimulated ERK2 activation. and shows that of SHP2 could not rescue the inhibitory of on ERK2 activation. Similarly, expression of the constitutively active not rescue the of on ERK2 activation that the activated SHP2 with Gab1 in to mediate ERK2 activation. To the that association of activated SHP2 PTPase with Gab1 is necessary for ERK2 activation, we a for expression of a protein in which is to the of shows that the Gab1FF-SHP2ΔN chimera was in cells. cells transfected with Gab1FF-SHP2ΔN constitutively ERK2 activity and and and that of were in the for the ERK2 activity The ERK2 activity in transfected cells was on the of Gab1FF-SHP2ΔN to cells a of for the constitutive ERK2 activity in cells transfected with Gab1FF-SHP2ΔN was the basal of constitutive ERK2 activity was to that in cells transfected with expression for the of is of the of and to activate ERK2 constitutively in cells T. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, P. Gutkind Nature. PubMed Scopus Google Scholar). in that phosphopeptides containing Tyr(P)-627 and Tyr(P)-659 are SHP2 To Tyr(P)-627 and Tyr(P)-659 of Gab1 are dephosphorylated by SHP2 in the we Gab1 with the or a inactive SHP2 catalytic to in cells. EGF stimulation for tyrosine of Gab1 and the of SHP2 in the Gab1 were in cells transfected with SHP2 and It has been that the catalytic to of can a with and these from H. Tonks N.K. Cell. Full Text PDF PubMed Scopus Google Scholar, T. M. M. H. S. A. PubMed Scopus Google Scholar, H. Neel B.G. J. Biol. Chem. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar). A and shows that tyrosine of Gab1 was in cells transfected with the SHP2. EGF stimulation resulted in a in Gab1 tyrosine that at The Gab1 tyrosine to of the at EGF stimulation in these cells. to the change in the of SHP2 in the Gab1 that Tyr(P)-627 Tyr(P)-659 of Gab1 were dephosphorylated in these cells. transfected with an of Gab1 tyrosine and Gab1-SHP2 in the A and EGF there was a in Gab1 tyrosine to a in the of SHP2 with Gab1 Therefore, was to Gab1 in a state in EGF-stimulated that Gab1 is a substrate of SHP2 in the cells. experiments have that Tyr-627 is required for SHP2 binding to Gab1 in cells with EGF or insulin T. L. J. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, S. S. J. M. Wong A.J. E. Mol. 1998; PubMed Scopus Google Scholar). The that is required for SHP2 binding to Gab1 in EGF-stimulated cells and for ERK2 activation by EGF. SHP2 contains two SH2 domains that are arranged in tandem in the N-terminal portion of SHP2, the of both Tyr-627 and Tyr-659 for Gab1-SHP2 association that both with the tandem SH2 domains of SHP2. is the Tyr(P)-627 and Tyr(P)-659 be to bind to SH2 domains in SHP2. Far Western blot binding that Tyr(P)-627 with the N-SH2 domain of SHP2, Tyr(P)-659 to the C-SH2 domain of SHP2. that it is for the tandem SH2 domains to bind to Tyr(P)-627 and Tyr(P)-659 in a specific orientation, in which Tyr(P)-627 binds to the N-SH2 domain and Tyr(P)-659 binds to the C-SH2 In other we assayed SHP2 PTPase activity We found that peptide but not peptide was sufficient to activate SHP2 PTPase in not SHP2 activation binding of a TAM to its N-SH2 domain, that has a affinity for the N-SH2 domain of SHP2. Therefore, of to the that Tyr(P)-627 binds to the N-SH2 domain while Tyr(P)-659 binds to the the specific binding of the tandem SH2 domains of SHP2 to the of IRS-1 has also been suggested S. Shoelson S.E. Neel B.G. J. Biol. Chem. Full Text PDF PubMed Google Scholar). structure analysis in the absence of N-SH2 domain binding the N-SH2 domain of SHP2 with its catalytic domain and thus the in an inactive state (3Hof P. Pluskey S. Dhe-Paganon S. Eck M.J. Shoelson S.E. Cell. 1998; 92: 441-450Abstract Full Text Full Text PDF PubMed Scopus (775) Google M.J. Pluskey S. Trub T. Harrison S.C. Shoelson S.E. Nature. 1996; 379: 277-280Crossref PubMed Scopus (171) Google Scholar). Binding of to the N-SH2 domain a change that activates the PTPase (3Hof P. Pluskey S. Dhe-Paganon S. Eck M.J. Shoelson S.E. Cell. 1998; 92: 441-450Abstract Full Text Full Text PDF PubMed Scopus (775) Google Scholar, 5Ottinger E.A. Botfield M.C. Shoelson S.E. J. Biol. Chem. 1998; 273: 729-735Abstract Full Text Full Text PDF PubMed Scopus (184) Google Scholar). We found that a bisphosphopeptide derived from Gab1 that contains both the Tyr(P)-627 and Tyr(P)-659 TAMs could activate SHP2 PTPase. In with IRS-1 were to the two TAMs that are by a amino (3Hof P. Pluskey S. Dhe-Paganon S. Eck M.J. Shoelson S.E. Cell. 1998; 92: 441-450Abstract Full Text Full Text PDF PubMed Scopus (775) Google Scholar, 5Ottinger E.A. Botfield M.C. Shoelson S.E. J. Biol. Chem. 1998; 273: 729-735Abstract Full Text Full Text PDF PubMed Scopus (184) Google Scholar). in the with a amino that has been for SHP2 activation. In a IRS-1 SHP2 activation was at of and and a activation of was (4Eck M.J. Pluskey S. Trub T. Harrison S.C. Shoelson S.E. Nature. 1996; 379: 277-280Crossref PubMed Scopus (171) Google Scholar). In a in was at the of that we have at and activates SHP2 and In to a low of the peptide to the substrate or we not to SHP2 activation. PTPase in these two to the in the that activates SHP2 PTPase that SHP2 binding to Gab1 in SHP2 activation, which is necessary for EGF-stimulated ERK2 activation. monophosphopeptides and could not activate SHP2 SHP2 was assayed peptide A mixture of and also could not activate SHP2 peptide were to SHP2. These that both Tyr-627 and Tyr-659 of Gab1 are required for EGF-stimulated ERK2 activation and that the phosphorylated Tyr-627 and Tyr-659 of Gab1 constitute a for the binding and activation of SHP2. these results suggest that the physical between the two TAMs in the Gab1 is necessary for SHP2 activation were its We found that expression of a constitutively active SHP2 was to the by of EGF-stimulated ERK2 activation. from with a deletion of the N-SH2 domain of SHP2 were defective in ERK activation W. G-S. J. Biol. Chem. 1998; 273: Full Text Full Text PDF PubMed Scopus (140) Google Scholar), that the activated SHP2 to with or its N-SH2 or both SH2 domains to mediate ERK activation. we found that expression of a protein of the constitutively active to the of the SHP2 binding defective resulted in a constitutive activation of ERK2 in cells. These results that the activated SHP2 with Gab1 in to mediate ERK activation by EGF and that constitutive association of activated SHP2 with Gab1 in a constitutively ERK2 kinase activity in the cells. The that the activated SHP2 has to with Gab1 in to mediate ERK activation is at but several SHP2 to a specific negative on SHP2 to a suggested M. M. G-S. Mol. Cell. Biol. PubMed Scopus Google SHP2 Gab1 to it to a specific to a target that is not with Phosphopeptides derived from and Tyr(P)-659 of Gab1 were dephosphorylated by SHP2 in vitro. substrate that Gab1 from with a in the of that was to Gab1 These experiments that Gab1 not only is a SHP2 activator in EGF-stimulated cells but also is a target of the activated SHP2 PTPase in the cells. We and for
Cunnick et al. (Fri,) studied this question.
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