Tyrosine 740 Phosphorylation of Discoidin Domain Receptor 2 by Src Stimulates Intramolecular Autophosphorylation and Shc Signaling Complex Formation

DDR2 is a receptor tyrosine kinase whose activating ligands are various collagens. DDR2-mediated cellular signaling has been shown to require Src activity. However, the precise mechanism underlying the Src dependence of DDR2 signaling is unknown. Here, using baculoviral co-expression of the DDR2 cytosolic domain and Src, we show that Src targets three tyrosine residues (Tyr-736, Tyr-740, and Tyr-741) in the activation loop of DDR2 for phosphorylation. This phosphorylation by Src stimulates DDR2 cis-autophosphorylation of additional tyrosine residues. In vitro Shc binding assays demonstrate that phosphotyrosines resulting from DDR2 autophosphorylation are involved in Shc binding to the DDR2 cytosolic domain. Mutating tyrosine 740 of DDR2 to phenylalanine stimulates autophosphorylation of DDR2 to an extent similar to that resulting from Src phosphorylation of DDR2. In addition, the DDR2 Y740F mutant protein displays collagen-independent, constitutively activated signaling. These findings suggest that tyrosine 740 inhibits DDR2 autophosphorylation. Collectively, our findings are consistent with the following mechanism for Src-dependent DDR2 activation and signaling: 1) ligand binding promotes phosphorylation of Tyr-740 in the DDR2 activation loop by Src; 2) Tyr-740 phosphorylation stimulates intramolecular autophosphorylation of DDR2; 3) DDR2 autophosphorylation generates cytosolic domain phosphotyrosines that promote the formation of DDR2 cytosolic domain-Shc signaling complexes. DDR2 is a receptor tyrosine kinase whose activating ligands are various collagens. DDR2-mediated cellular signaling has been shown to require Src activity. However, the precise mechanism underlying the Src dependence of DDR2 signaling is unknown. Here, using baculoviral co-expression of the DDR2 cytosolic domain and Src, we show that Src targets three tyrosine residues (Tyr-736, Tyr-740, and Tyr-741) in the activation loop of DDR2 for phosphorylation. This phosphorylation by Src stimulates DDR2 cis-autophosphorylation of additional tyrosine residues. In vitro Shc binding assays demonstrate that phosphotyrosines resulting from DDR2 autophosphorylation are involved in Shc binding to the DDR2 cytosolic domain. Mutating tyrosine 740 of DDR2 to phenylalanine stimulates autophosphorylation of DDR2 to an extent similar to that resulting from Src phosphorylation of DDR2. In addition, the DDR2 Y740F mutant protein displays collagen-independent, constitutively activated signaling. These findings suggest that tyrosine 740 inhibits DDR2 autophosphorylation. Collectively, our findings are consistent with the following mechanism for Src-dependent DDR2 activation and signaling: 1) ligand binding promotes phosphorylation of Tyr-740 in the DDR2 activation loop by Src; 2) Tyr-740 phosphorylation stimulates intramolecular autophosphorylation of DDR2; 3) DDR2 autophosphorylation generates cytosolic domain phosphotyrosines that promote the formation of DDR2 cytosolic domain-Shc signaling complexes. The discoidin domain receptor (DDR) 2The abbreviations used are:DDRdiscoidin domain receptorDDR2 CKDdiscoidin domain receptor 2 cytosolic tyrosine kinase domainkd-DDR2kinase defective discoidin domain receptor 2GSTglutathione S-transferaseMMPmatrix metalloproteinaseIRTKinsulin receptor tyrosine kinaseHSChepatic stellate cellCKDcytosolic kinase domainCMVcytomegalovirusTBSTris-buffered salineγ-S-ATPγ-S-adenosine triphosphate 2The abbreviations used are:DDRdiscoidin domain receptorDDR2 CKDdiscoidin domain receptor 2 cytosolic tyrosine kinase domainkd-DDR2kinase defective discoidin domain receptor 2GSTglutathione S-transferaseMMPmatrix metalloproteinaseIRTKinsulin receptor tyrosine kinaseHSChepatic stellate cellCKDcytosolic kinase domainCMVcytomegalovirusTBSTris-buffered salineγ-S-ATPγ-S-adenosine triphosphate family, including DDR1 and DDR2, belongs to receptor tyrosine kinases (RTKs) family. Its extracellular part, containing the so-called discoidin domain, binds to various collagen proteins as their activating ligands (1Shrivastava A. Radziejewski C. Campbell E. Kovac L. McGlynn M. Ryan T.E. Davis S. Goldfarb M.P. Glass D.J. Lemke G. Yancopoulos G.D. Mol. Cell. 1997; 1: 25-34Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar, 2Vogel W. Gish G.D. Alves F. Pawson T. Mol. Cell. 1997; 1: 13-23Abstract Full Text Full Text PDF PubMed Scopus (766) Google Scholar, 3Vogel W. Brakebusch C. Fassler R. Alves F. Ruggiero F. Pawson T. J. Biol. Chem. 2000; 275: 5779-5784Abstract Full Text Full Text PDF PubMed Scopus (123) Google Scholar), and its intracellular part possesses a domain of tyrosine kinase, which shares ∼50% sequence homology with that of the Trk family of neurotrophin receptors as well as with insulin receptor (4Lai C. Lemke G. Oncogene. 1994; 9: 877-883PubMed Google Scholar, 5Laval S. Butler R. Shelling A.N. Hanby A.M. Poulsom R. Ganesan T.S. Cell Growth & Differ. 1994; 5: 1173-1183PubMed Google Scholar, 6Ebina Y. Ellis L. Jarnagin K. Edery M. Graf L. Clauser E. Ou J. Masiarz F. Kan Y.W. Goldfine I.D. Roth R.A. Cell. 1985; 40: 747-758Abstract Full Text PDF PubMed Scopus (957) Google Scholar). discoidin domain receptor discoidin domain receptor 2 cytosolic tyrosine kinase domain kinase defective discoidin domain receptor 2 glutathione S-transferase matrix metalloproteinase insulin receptor tyrosine kinase hepatic stellate cell cytosolic kinase domain cytomegalovirus Tris-buffered saline γ-S-adenosine triphosphate discoidin domain receptor discoidin domain receptor 2 cytosolic tyrosine kinase domain kinase defective discoidin domain receptor 2 glutathione S-transferase matrix metalloproteinase insulin receptor tyrosine kinase hepatic stellate cell cytosolic kinase domain cytomegalovirus Tris-buffered saline γ-S-adenosine triphosphate Involvement of DDR proteins in the proliferation of various cell types has been reported. Increased DDR1 expression is observed in keratinocytes of the skin and smooth muscle cells around blood vessels when the tissues are injured (7Ferri N. Carragher N.O. Raines E.W. Am. J. Pathol. 2004; 164: 1575-1585Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar, 8Hou G. Vogel W.F. Bendeck M.P. Circ. Res. 2002; 90: 1147-1149Crossref PubMed Scopus (124) Google Scholar). DDR1 is also expressed in monocyte-derived cells where it is believed to play a role in collagen binding and cell differentiation (9Matsuyama W. Faure M. Yoshimura T. J. Immunol. 2003; 171: 3520-3532Crossref PubMed Scopus (45) Google Scholar, 10Matsuyama W. Kamohara H. Galligan C. Faure M. Yoshimura T. FASEB J. 2003; 17: 1286-1288Crossref PubMed Scopus (43) Google Scholar). DDR2 expression is observed in mesenchymal cells and is involved in bone growth (11Islam S. Kermode T. Sultana D. Moskowitz R.W. Mukhtar H. Malemud C.J. Goldberg V.M. Haqqi T.M. Osteoarthritis Cartilage. 2001; 9: 684-693Abstract Full Text PDF PubMed Scopus (30) Google Scholar). During liver fibrosis, induction and activation of DDR2 occur in liver stellate cells, and its tyrosine kinase activity is necessary for the proliferation of stellate cells and for the increase of collagen and MMP-2 synthesis (12Olaso E. Ikeda K. Eng F.J. Xu L. Wang L.H. Lin H.C. Friedman S.L. J. Clin. Invest. 2001; 108: 1369-1378Crossref PubMed Scopus (243) Google Scholar, 13Olaso E. Labrador J.P. Wang L. Ikeda K. Eng F.J. Klein R. Lovett D.H. Lin H.C. Friedman S.L. J. Biol. Chem. 2002; 277: 3606-3613Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar). In rheumatoid arthritis, DDR2 induction is also observed in activated synovial fibroblasts and is thought to stimulate the growth of these cells and MMP-1 synthesis (14Wang J. Lu H. Liu X. Deng Y. Sun T. Li F. Ji S. Nie X. Yao L. J. Autoimmun. 2002; 19: 161-168Crossref PubMed Scopus (31) Google Scholar). In addition, the induction of DDR proteins is implicated in breast and ovarian cancer, and is correlated with metastasis (15Heinzelmann-Schwarz V.A. Gardiner-Garden M. Henshall S.M. Scurry J. Scolyer R.A. Davies M.J. Heinzelmann M. Kalish L.H. Bali A. Kench J.G. Edwards L.S. Vanden Bergh P.M. Hacker N.F. Sutherland R.L. O'Brien P.M. Clin. Cancer Res. 2004; 10: 4427-4436Crossref PubMed Scopus (176) Google Scholar, 16Evtimova V. Zeillinger R. Weidle U.H. Tumour Biol. 2003; 24: 189-198Crossref PubMed Scopus (33) Google Scholar). Autophosphorylation of the cytosolic domain of RTKs is typically a critical event for the activation of RTK-mediated cellular signaling (17Hubbard S.R. Till J.H. Annu. Rev. Biochem. 2000; 69: 373-398Crossref PubMed Scopus (859) Google Scholar). Three-dimensional structure determination as well as associated biochemical and genetic analysis of several RTKs have helped to reveal the detailed molecular mechanisms of RTK autophosphorylation (18Hubbard S.R. Wei L. Ellis L. Hendrickson W.A. Nature. 1994; 372: 746-754Crossref PubMed Scopus (948) Google Scholar, 19Hubbard S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar, 20Mohammadi M. Schlessinger J. Hubbard S.R. Cell. 1996; 86: 577-587Abstract Full Text Full Text PDF PubMed Scopus (330) Google Scholar, 25Wei L. Hubbard S.R. Hendrickson W.A. Ellis L. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar, K. J. Biol. Chem. Full Text PDF PubMed Google Scholar). the insulin and growth ligand binding the of three tyrosine residues in a so-called activation loop and of the kinase This phosphorylation a in the structure of the activation loop and stimulates RTK activity by and to the (17Hubbard S.R. Till J.H. Annu. Rev. Biochem. 2000; 69: 373-398Crossref PubMed Scopus (859) Google Scholar, 19Hubbard S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar). autophosphorylation of tyrosine residues in the and by the kinase activity of these receptors (17Hubbard S.R. Till J.H. Annu. Rev. Biochem. 2000; 69: 373-398Crossref PubMed Scopus (859) Google Scholar, J. Biol. Chem. 1994; Full Text PDF PubMed Google Scholar, R.A. 1997; PubMed Scopus Google Scholar). of cells with the ligand collagen tyrosine phosphorylation of the cytosolic domain of DDR1 and DDR2 (1Shrivastava A. Radziejewski C. Campbell E. Kovac L. McGlynn M. Ryan T.E. Davis S. Goldfarb M.P. Glass D.J. Lemke G. Yancopoulos G.D. Mol. Cell. 1997; 1: 25-34Abstract Full Text Full Text PDF PubMed Scopus (431) Google Scholar, 2Vogel W. Gish G.D. Alves F. Pawson T. Mol. Cell. 1997; 1: 13-23Abstract Full Text Full Text PDF PubMed Scopus (766) Google Scholar). activity the of tyrosine phosphorylation in DDR proteins ligand In tyrosine phosphorylation of RTKs the of ligand The for is In it has been that DDR2-mediated signaling Src tyrosine kinase activity K. Wang L.H. R. H. E. Eng F.J. Labrador Klein R. Lovett D. Yancopoulos G.D. Friedman S.L. Lin H.C. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). In we the precise mechanism for activation of DDR2 cytosolic signaling using in vitro baculoviral expression using glutathione S-transferase of expressed the to the by a of from a the and the in the baculoviral expression of DDR2 cytosolic kinase domain a DDR2 from the of expression of DDR1 cytosolic kinase domain from and by a of and from the of and the of to their baculoviral expression expression DDR2 the of the by of and of DDR2 by with The MMP-1 from to from from cells and the of to MMP-1 The sequence of by in by to a baculoviral expression of DDR2 kinase domain to a cytosolic domain by DDR2 cytosolic domain with a to the of from using of containing an containing a containing a and containing a to the three tyrosine residues and Tyr-741) in the activation of DDR2 tyrosine kinase domain DDR2 tyrosine kinase domain, we from DDR2 and from DDR2 using of of containing and of containing as well as a as Y740F and of the in The and Y740F the DDR2 expression to and expression by the sequence with the containing from the baculoviral expression Cell and cells in with and 2 and cells using and cells using in the of and cells by S. L. Friedman of and of and of baculoviral expression cells with from a from to the The to a of cells with of and for DDR2 kinase domain proteins using a and by cells from in of of a and by for The for using a and the to of a with and with the The proteins with of containing glutathione and to using The to a with a of containing and and with the of using a The kinase domain protein and using a of proteins to glutathione cells in of the by and using for of of glutathione to the and the for the by with for three Autophosphorylation and of the autophosphorylation activity of DDR2, the using of DDR2 kinase domain protein in of containing and of the by a of with for 2 The in the of containing and the using and The in the DDR2 kinase protein by and using a the autophosphorylation from to DDR2 cytosolic the in the and DDR2 by in the by and the of from the of the as K. Mol. 2003; PubMed Scopus Google Scholar). of DDR2 tyrosine kinase activity using of the kinase in of containing of as and of the by a of used as a the as a The with for and the of and using a and and DDR2 from Cell and and Shc from for 2 in and in to and by in for The with for 2 and with with for and protein by Shc a of Shc of cells the of the 2 cells and by in of of and a The by for and used for Shc binding assays as proteins to glutathione as the of to kinase containing and of to the glutathione for and three with autophosphorylation using of Shc cell to the and the for the with for three and the proteins the by in for 2 The proteins by using to Shc MMP-1 and of cells and with of of MMP-1 with of well using to the cells with collagen cells and activity cell DDR2 of expression cells in a with of using cells to and to with a increase of from to cell for the expression of DDR2 by using the to DDR2. MMP-2 cells of as well as cells of and in and for The from well and for the of MMP-2 using an from to the Src the DDR2 we expressed the DDR2 tyrosine kinase domain as a in cells by baculoviral the expressed tyrosine phosphorylation in cells as shown by using However, when with tyrosine kinase in cells, induction of tyrosine phosphorylation including tyrosine phosphorylation in the that tyrosine phosphorylation the DDR2 tyrosine kinase domain. In co-expression of with a Src tyrosine phosphorylation of These that Src the kinase domain of DDR2. in of these similar of used as shown by and the expression of Src proteins in cells by phosphorylation by Src the DDR2 kinase domain including the kinase domain and of of these kinases with Src in cells, Src-dependent tyrosine phosphorylation of and and phosphorylation of Src-dependent tyrosine phosphorylation of of kinase used in these and the of protein by with the tyrosine phosphorylation we Src co-expression to Src a with a mutant of the DDR2 kinase domain This mutant by a in the to This the tyrosine kinase activity of the protein co-expression with Src in cells, the protein tyrosine phosphorylation tyrosine phosphorylation of This that tyrosine phosphorylation of the DDR2 tyrosine kinase domain by Src and in stimulates autophosphorylation of the DDR2 tyrosine kinase domain. The the protein in an in vitro phosphorylation as well This is consistent with the that Src the DDR2 tyrosine kinase domain in our co-expression by Src the Autophosphorylation of DDR2 and Its the of the observed DDR2 phosphorylation by Src, we stimulates the autophosphorylation activity of DDR2. by Src cells with Src and In when the of Src to baculoviral the of tyrosine phosphorylation in a of we that of tyrosine phosphorylation by Src by the in a of with a of of in to of using and a Src protein in the as by from cells in the 2 of with the the autophosphorylation of Src DDR2 and DDR2 in kinase containing for for autophosphorylation and autophosphorylation In the autophosphorylation activity of and to that of These suggest that phosphorylation by Src a that inhibits DDR2 kinase domain autophosphorylation. we the of Src phosphorylation the activity of the DDR2 kinase the and that the DDR2 kinase by Src has to activity these to that of the DDR2 kinase we autophosphorylation the DDR2 kinase domain its tyrosine kinase activity. DDR2 kinase domain proteins with of tyrosine phosphorylation by in vitro autophosphorylation for in the of DDR2 kinase domain co-expression with Src in cells used for the proteins in tyrosine phosphorylation as by tyrosine kinase activity This that tyrosine phosphorylation of DDR2 by Src, DDR2 the of DDR2 tyrosine kinase activity. we the of and using as a tyrosine phosphorylation of the DDR2 kinase by Src the of DDR2 for from to and by This that Src phosphorylation of the DDR2 cytosolic kinase domain its for as well as its Autophosphorylation of the DDR2 an that phosphorylation by Src and stimulates autophosphorylation of the DDR2 receptor tyrosine kinase domain. In an to the autophosphorylation in we DDR2 autophosphorylation cis-autophosphorylation we the autophosphorylation of from to DDR2 in the autophosphorylation as the of from to the of of of as the and with a of the by to The by in with the a of molecular in the of This that autophosphorylation of DDR2 cytosolic domain an in the DDR2 the of Src activation mechanism of has been well (18Hubbard S.R. Wei L. Ellis L. Hendrickson W.A. Nature. 1994; 372: 746-754Crossref PubMed Scopus (948) Google Scholar, 19Hubbard S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar, R.A. 1997; PubMed Scopus Google Scholar, 25Wei L. Hubbard S.R. Hendrickson W.A. Ellis L. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar, K. J. Biol. Chem. Full Text PDF PubMed Google Scholar). These have that phosphorylation of three tyrosine residues and the activation loop of is critical for the activation of autophosphorylation as well as for its tyrosine kinase activity S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar, 25Wei L. Hubbard S.R. Hendrickson W.A. Ellis L. J. Biol. Chem. 1995; 270: Full Text Full Text PDF PubMed Scopus Google Scholar, K. J. Biol. Chem. Full Text PDF PubMed Google Scholar). that three tyrosine Tyr-740, and of DDR2, are to involved in the activation of DDR2 kinase activity by Src the kinase domain of DDR2 is to that of the insulin and we that the three tyrosine residues to and in the activation loop of by the kinase domain of DDR2 and the insulin receptor the of DDR2 activation loop tyrosine phosphorylation by Src, we in in and in the three residues in the of baculoviral the DDR2 mutant These are in The to DDR2 autophosphorylation and DDR2 tyrosine phosphorylation from Src activity. we expressed the in cells, we DDR2 expression from of the and co-expression with Src in cells, the of Src phosphorylation of these mutant proteins by using a phosphorylation in the The and tyrosine phosphorylation observed for DDR2 with activation loop These that Src three activation loop tyrosine residues of DDR2. of 740 to in the of DDR2 the of Src DDR2 the of Tyr-740, and in DDR2 we the baculoviral expression of The and its mutant proteins expressed in cells, with Src and by with a in the of Src mutant proteins containing phenylalanine 740 and tyrosine phosphorylation a to that of the DDR2 with Src In the and of Src with mutant DDR2 proteins Y740F increase the of DDR2 tyrosine phosphorylation the observed in the of Src co-expression These suggest that tyrosine 740 with phenylalanine in of the of Src phosphorylation of the DDR2 kinase domain. In addition, we that phosphorylation tyrosine residues in the DDR2 activation loop in these the mutant a increase in tyrosine phosphorylation we the Y740F stimulate autophosphorylation of the DDR2 tyrosine kinase in the in vitro autophosphorylation of and various DDR2 mutant mutant proteins the Y740F of by tyrosine autophosphorylation to DDR2 and mutant proteins that the Tyr-740 These that the Y740F stimulates DDR2 kinase autophosphorylation Src phosphorylation of DDR2. that tyrosine 740 DDR2 as for DDR2, we observed that autophosphorylation of the DDR2 Y740F mutant an intramolecular mechanism DDR2 Autophosphorylation for Shc to the DDR2 has been shown that the domain signaling protein Shc binds to activated DDR2 receptors in the liver hepatic stellate cell K. Wang L.H. R. H. E. Eng F.J. Labrador Klein R. Lovett D. Yancopoulos G.D. Friedman S.L. Lin H.C. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar). Shc binds to the DDR2 cytosolic domain to DDR2 that has Src phosphorylation and autophosphorylation. cytosolic from cells with and Src used for in vitro Shc binding assays in cell DDR2 proteins to autophosphorylation for autophosphorylation DDR2 expressed Src co-expression to of to autophosphorylation to Shc binding In DDR2 with Src Shc and binding by the in vitro autophosphorylation also shown to increase DDR2 tyrosine This that autophosphorylation of the DDR2 cytosolic domain is necessary for Shc we the Shc binding with Y740F and mutant in that the Y740F and autophosphorylation activity by In Shc binding assays with the autophosphorylation Y740F and binding similar to that of DDR2 cytosolic domain by Src and to autophosphorylation This our that the Y740F the of Src with to DDR2 autophosphorylation and Shc binding and the that Src DDR2 signaling by The that the mutant DDR2 protein Shc binding autophosphorylation that the three tyrosine residues in the activation loop of DDR2 in Shc binding and that tyrosine residues in DDR2 are involved in Shc in the DDR2 has been implicated in Shc binding K. Wang L.H. R. H. E. Eng F.J. Labrador Klein R. Lovett D. Yancopoulos G.D. Friedman S.L. Lin H.C. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar), we a DDR2 Shc with the the mutant is by Src in cells, and phosphorylation its autophosphorylation activity that DDR2 tyrosine residues the that is of residues involved in Shc observed the in vitro autophosphorylation tyrosine phosphorylation of the mutant protein by Src, the Shc binding to the DDR2 cytosolic domain This the that is involved in Shc In addition, our suggest that autophosphorylation is for Shc DDR2 Y740F MMP-1 and MMP-2 of are consistent with the that the DDR2 Y740F DDR2 signaling in a it shown that collagen to cells DDR2 the expression of MMP-1 and MMP-2 W. Gish G.D. Alves F. Pawson T. Mol. Cell. 1997; 1: 13-23Abstract Full Text Full Text PDF PubMed Scopus (766) Google Scholar, E. Ikeda K. Eng F.J. Xu L. Wang L.H. Lin H.C. Friedman S.L. J. Clin. Invest. 2001; 108: 1369-1378Crossref PubMed Scopus (243) Google Scholar, J. Lu H. Liu X. Deng Y. Sun T. Li F. Ji S. Nie X. Yao L. J. Autoimmun. 2002; 19: 161-168Crossref PubMed Scopus (31) Google Scholar). we expression of DDR2 Y740F MMP-1 and MMP-2 expression in a we MMP-1 in cells a of the DDR2, the Y740F the of the MMP-1 to to to signaling A. K. K. G. 2003; PubMed Scopus (31) Google Scholar). the cell with the ligand that MMP-1 activity by collagen in cells with the DDR2 expression However, in the of mutant the MMP-1 activity that of DDR2 and with collagen as we activation of MMP-1 activity by collagen in of DDR2 the activation of DDR2 signaling by Y740F we cells from cells that DDR2 DDR2 and DDR2 Y740F DDR2 and in to the of DDR2 protein cells MMP-2 expression in the three cells DDR2 Y740F in the three of DDR2 by an of the when cells the to the induction of MMP-2 we a increase of MMP-2 expression the observed in in the of the three cells DDR2 an induction in three cells DDR2. cells to the cells DDR2 Collectively, these the that the Y740F mutant constitutively signaling and are consistent with the of our biochemical In we used co-expression of Src, the DDR2 kinase domain, and a of DDR2 mutant proteins to the mechanism of DDR2 receptor tyrosine kinase signaling. This to of the associated with Src phosphorylation of DDR2 in phosphorylation in vitro with DDR2 kinase domain and Src proteins of DDR2 tyrosine phosphorylation. In addition, the tyrosine kinase activity of DDR2 is analysis is to a of DDR2 activity in vitro phosphorylation we that DDR2 tyrosine kinase of its activity J. H. H. J. Y. and Here, we that DDR2 tyrosine kinase the phosphorylation the tyrosine residues of its activation loop by Src to show a autophosphorylation activity in our Src tyrosine kinase activity also to signaling by growth growth and T. Cell Biol. 1995; PubMed Scopus Google Scholar, S. A. PubMed Scopus Google Scholar, Y. J. Biol. Chem. 1985; Full Text PDF PubMed Google Scholar). However, when we the autophosphorylation activity of growth receptor and kinase from cells, autophosphorylation activity in the of Src and activity when co-expression with J. H. H. J. Y. and has been that the activated with three tyrosine residues in its activation loop by in vitro autophosphorylation using protein from expressed cells S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar). These suggest that growth receptor and Src activity for their Src kinase activity is for the activation of the DDR2 tyrosine In we have also in the molecular mechanism underlying the and Shc binding of the DDR2 cytosolic domain. the three activation loop Tyr-740 is for the autophosphorylation activity of DDR2. we observed that DDR2 by Src and the Y740F mutant in of autophosphorylation and Shc our findings that the of Src in the activation of DDR2 receptor signaling is the induction of phosphorylation tyrosine is the three tyrosine residues in the activation loop by autophosphorylation from by However, it is that tyrosine residues in DDR2 in the autophosphorylation and of these residues is for the binding of Shc that tyrosine in the of DDR2 by Src and that is the for Shc binding K. Wang L.H. R. H. E. Eng F.J. Labrador Klein R. Lovett D. Yancopoulos G.D. Friedman S.L. Lin H.C. J. Biol. Chem. 2002; 277: Full Text Full Text PDF PubMed Scopus Google Scholar), our that phosphorylation occur by Src, it from autophosphorylation. our the that involved in Shc In receptor tyrosine phosphorylation of tyrosine residues in the activation loop is a event in the of RTK autophosphorylation activity (17Hubbard S.R. Till J.H. Annu. Rev. Biochem. 2000; 69: 373-398Crossref PubMed Scopus (859) Google Scholar). In it shown that to Tyr-740 of of a with the to the activation loop an S.R. EMBO J. 1997; 16: 5572-5581Crossref PubMed Scopus (768) Google Scholar). to our it that an tyrosine 740 in the DDR2 activation loop also has a critical role for DDR2 tyrosine kinase activity. Its to phenylalanine DDR2 tyrosine kinase from a of to an to the intramolecular of DDR2 is as a of various including fibrosis, rheumatoid and and (12Olaso E. Ikeda K. Eng F.J. Xu L. Wang L.H. Lin H.C. Friedman S.L. J. Clin. Invest. 2001; 108: 1369-1378Crossref PubMed Scopus (243) Google Scholar, 13Olaso E. Labrador J.P. Wang L. Ikeda K. Eng F.J. Klein R. Lovett D.H. Lin H.C. Friedman S.L. J. Biol. Chem. 2002; 277: 3606-3613Abstract Full Text Full Text PDF PubMed Scopus (193) Google Scholar, J. Lu H. Liu X. Deng Y. Sun T. Li F. Ji S. Nie X. Yao L. J. Autoimmun. 2002; 19: 161-168Crossref PubMed Scopus (31) Google Scholar, V.A. Gardiner-Garden M. Henshall S.M. Scurry J. Scolyer R.A. Davies M.J. Heinzelmann M. Kalish L.H. Bali A. Kench J.G. Edwards L.S. Vanden Bergh P.M. Hacker N.F. Sutherland R.L. O'Brien P.M. Clin. Cancer Res. 2004; 10: 4427-4436Crossref PubMed Scopus (176) Google Scholar, 16Evtimova V. Zeillinger R. Weidle U.H. Tumour Biol. 2003; 24: 189-198Crossref PubMed Scopus (33) Google Scholar, L. H. D. K. Li Y. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar). to the precise mechanism of DDR2 tyrosine kinase activation and its signaling for the of and DDR2 for these suggest that the structure around the of the DDR2 tyrosine kinase its activation by the for and are and activated DDR2 tyrosine In it to that the binding of the activated DDR2 tyrosine

Récupéré en direct depuis OpenAlex et désinversé. Les résumés ne sont pas conservés dans cette base de données : les index inversés représentent 8,6 Go des 9,3 Go de texte de la base, et le serveur dispose de 13 Go libres.