Work

M-CSF Regulates the Cytoskeleton via Recruitment of a Multimeric Signaling Complex to c-Fms Tyr-559/697/721

Year: 2007

Type: article

Abstract: M-CSF is known to induce cytoskeletal reorganization in macrophages and osteoclasts by activation of phosphatidylinositol 3-kinase (PI3K) and c-Src, but the detailed mechanisms remain unclear. We find, unexpectedly, that tyrosine (Tyr) to phenylalanine (Phe) mutation of Tyr-721, the PI3K binding site in the M-CSF receptor c-Fms, fails to suppress cytoskeletal remodeling or actin ring formation. In contrast, mutation of c-Fms Tyr-559 to Phe blocks M-CSF-induced cytoskeletal reorganization by inhibiting formation of a Src Family Kinase SFK·c-Cbl·PI3K complex and the downstream activation of Vav3 and Rac, two key mediators of actin remodeling. Using an add-back approach in which specific Tyr residues are reinserted into c-Fms inactivated by the absence of all seven functionally important Tyr residues, we find that Tyr-559 is necessary but not sufficient to transduce M-CSF-dependent cytoskeletal reorganization. Furthermore, this same add-back approach identifies important roles for Tyr-697 and Tyr-721 in collaborating with Tyr-559 to recruit a multimeric signaling complex that can transduce signals from c-Fms to the actin cytoskeleton. M-CSF is known to induce cytoskeletal reorganization in macrophages and osteoclasts by activation of phosphatidylinositol 3-kinase (PI3K) and c-Src, but the detailed mechanisms remain unclear. We find, unexpectedly, that tyrosine (Tyr) to phenylalanine (Phe) mutation of Tyr-721, the PI3K binding site in the M-CSF receptor c-Fms, fails to suppress cytoskeletal remodeling or actin ring formation. In contrast, mutation of c-Fms Tyr-559 to Phe blocks M-CSF-induced cytoskeletal reorganization by inhibiting formation of a Src Family Kinase SFK·c-Cbl·PI3K complex and the downstream activation of Vav3 and Rac, two key mediators of actin remodeling. Using an add-back approach in which specific Tyr residues are reinserted into c-Fms inactivated by the absence of all seven functionally important Tyr residues, we find that Tyr-559 is necessary but not sufficient to transduce M-CSF-dependent cytoskeletal reorganization. Furthermore, this same add-back approach identifies important roles for Tyr-697 and Tyr-721 in collaborating with Tyr-559 to recruit a multimeric signaling complex that can transduce signals from c-Fms to the actin cytoskeleton. Cell motility is a dynamic process consisting of repeated cycles of protrusion of filopodia and lamellipodia at the leading front, adhesion of the protruding edge to the substratum via focal complexes, contraction of cytoplasmic actomyosin, and finally release from contact sites at the tail of the cell (1Lauffenburger D.A. Horwitz A.F. Cell. 1996; 84: 359-369Abstract Full Text Full Text PDF PubMed Scopus (3274) Google Scholar, 2Webb D.J. Parsons J.T. Horwitz A.F. Nat. Cell Biol. 2002; 4: E97-E100Crossref PubMed Scopus (602) Google Scholar, 3Ridley A.J. Schwartz M.A. Burridge K. Firtel R.A. Ginsberg M.H. Borisy G. Parsons J.T. Horwitz A.R. Science. 2003; 302: 1704-1709Crossref PubMed Scopus (3813) Google Scholar). The locomotory apparatus of most cells works against cell-to-substratum adhesions. In macrophages and osteoclasts (OCs), 2The abbreviations used are: OC, osteoclast; RTK, receptor tyrosine kinase; WT, wild type; SFK, Src family kinase; PI3K, phosphatidylinositol 3-kinase; Epo, erythropoietin; EpoR, Epo receptor; RANKL, receptor activator of NFκB ligand; BMM, bone marrow-derived macrophage; Ab, antibody; ERK, extracellular signal-regulated kinase. 2The abbreviations used are: OC, osteoclast; RTK, receptor tyrosine kinase; WT, wild type; SFK, Src family kinase; PI3K, phosphatidylinositol 3-kinase; Epo, erythropoietin; EpoR, Epo receptor; RANKL, receptor activator of NFκB ligand; BMM, bone marrow-derived macrophage; Ab, antibody; ERK, extracellular signal-regulated kinase. their differentiated progeny, integrin-mediated contacts to the substratum, are called podosomes (4Gavazzi I. Nermut M.V. Marchisio P.C. J. Cell Sci. 1989; 94: 85-89PubMed Google Scholar, 5Pfaff M. Jurdic P. J. Cell Sci. 2001; 114: 2775-2786Crossref PubMed Google Scholar). In contrast to the more stable focal adhesions, podosomes are very dynamic structures that assemble and disassemble rapidly (6Stickel S. Wang Y. J. Cell Biol. 1987; 104: 1521-1526Crossref PubMed Scopus (45) Google Scholar, 7Destaing O. Saltel F. Gilquin B. Chabadel A. Khochbin S. Ory S. Jurdic P. J. Cell Sci. 2005; 118: 2901-2911Crossref PubMed Scopus (188) Google Scholar), reflecting the motile nature of OCs and their myeloid precursors. In migrating macrophages and OCs, podosomes are confined to the periphery of the cell, where their reorganization results in lamellipodia formation, a process dependent on de novo actin assembly by the Arp2/3-Ena·Wiskott-Aldrich syndrome protein complex (8Nakagawa H. Miki H. Ito M. Ohashi K. Takenawa T. Miyamoto S. J. Cell Sci. 2001; 114: 1555-1565Crossref PubMed Google Scholar). Activation of actin polymerization is regulated by Rac GTPase acting through one or more members of the Wiskott-Aldrich syndrome protein family (9Takenawa T. Miki H. J. Cell Sci. 2001; 114: 1801-1809Crossref PubMed Google Scholar, 10Bi E. Zigmond S.H. Curr. Biol. 1999; 9: R160-R163Abstract Full Text Full Text PDF PubMed Scopus (58) Google Scholar). The fact that podosome formation, macrophage chemotaxis, and osteoclast movement on bone are impaired by mutation of the Wiskott-Aldrich syndrome protein (11Calle Y. Jones G.E. Jagger C. Fuller K. Blundell M.P. Chow J. Chambers T. Thrasher A.J. Blood. 2004; 103: 3552-3561Crossref PubMed Scopus (98) Google Scholar), inhibition of Rac activity (12Faccio R. Novack D.V. Zallone A. Ross F.P. Teitelbaum S.L. J. Cell Biol. 2003; 162: 499-509Crossref PubMed Scopus (149) Google Scholar), or small interfering RNA suppression of the actin-related protein 2/3 complex (13Razzouk S. Lieberherr M. Cournot G. Eur. J. Cell Biol. 1999; 78: 249-255Crossref PubMed Scopus (70) Google Scholar, 14Hurst I.R. Zuo J. Jiang J. Holliday L.S. J. Bone Miner. Res. 2004; 19: 499-506Crossref PubMed Scopus (69) Google Scholar) attests to the importance of actin assembly in macrophages and OC function. Macrophages and OCs both respond to chemotactic stimuli initiated by M-CSF. Re-addition of M-CSF to quiescent BAC1.2F5 macrophages stimulates rapid cytoskeletal reorganization and cell motility within a few minutes, followed by chemotactic migration up a gradient of diffusing cytokine (15Boocock C.A. Jones G.E. Stanley E.R. Pollard J.W. J. Cell Sci. 1989; 93: 447-456PubMed Google Scholar). Exposure to the cytokine also increases cell spreading, membrane ruffling, and motility of osteoclasts (12Faccio R. Novack D.V. Zallone A. Ross F.P. Teitelbaum S.L. J. Cell Biol. 2003; 162: 499-509Crossref PubMed Scopus (149) Google Scholar, 16Wells C.M. Walmsley M. Ooi S. Tybulewicz V. Ridley A.J. J. Cell Sci. 2004; 117: 1259-1268Crossref PubMed Scopus (153) Google Scholar) and macrophages (16Wells C.M. Walmsley M. Ooi S. Tybulewicz V. Ridley A.J. J. Cell Sci. 2004; 117: 1259-1268Crossref PubMed Scopus (153) Google Scholar). M-CSF binds to its transmembrane receptor tyrosine kinase (RTK) c-Fms and induces receptor autophosphorylation at seven tyrosine residues within the cytoplasmic domain (17Pixley F.J. Stanley E.R. Trends Cell Biol. 2004; 14: 628-638Abstract Full Text Full Text PDF PubMed Scopus (608) Google Scholar). Several Src homology 2 domain-containing molecules are recruited to the phospho-Tyr residues upon M-CSF binding and initiate different signaling cascades that lead to cell proliferation, differentiation, or cytoskeletal reorganization (17Pixley F.J. Stanley E.R. Trends Cell Biol. 2004; 14: 628-638Abstract Full Text Full Text PDF PubMed Scopus (608) Google Scholar). The identity of the molecules binding the cytoplasmic domain of c-Fms has been determined by expressing wild-type (WT) and tyrosine (Tyr) to phenylalanine (Phe) mutated forms of the RTK in various cell lines. The resulting model suggests that c-Fms Tyr-559 and Tyr-721 bind Src Family Kinases (SFKs) and phosphatidylinositol 3-kinase (PI3K), respectively, Tyr-697 and is a binding site for The Scholar, J. Biol. PubMed Scopus Google Scholar, PubMed Scopus Google Scholar). the detailed signaling by M-CSF is not PI3K and are recruited to c-Fms and migration of macrophages and of or macrophages is impaired in to the cytokine G.E. E. R. C. E. M.P. Ridley A.J. Cell Res. 2003; PubMed Scopus Google Scholar, V. J. D.A. R. Blood. 2005; PubMed Scopus Google Scholar). a PI3K OC migration M-CSF and blocks bone in and in J. Bone Miner. Res. PubMed Scopus Google Scholar, I. T. S. H. K. Y. T. T. PubMed Scopus Google Scholar). OCs to M-CSF impaired migration and a of cytoskeletal reorganization M. S. M. R. J. PubMed Scopus Google Scholar), that to their in activity P. C. R. A. Cell. Full Text PDF PubMed Scopus Google Scholar). the myeloid cell which into M-CSF-dependent into more macrophages a specific c-Fms tyrosine that the to recruit Src family the importance of c-Fms Tyr-559 and Src signaling C.M. J. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). The of this is to the signaling initiated by M-CSF that the cytoskeletal in macrophages and OCs, of which this we cells with a receptor consisting of the extracellular domain of receptor and the transmembrane and cytoplasmic of c-Fms, in which the functionally tyrosine residues or mutated to or in We this model in that Epo of cells expressing the into OCs to and M-CSF S. S. Teitelbaum S.L. Ross F.P. 2002; PubMed Scopus Google Scholar). We that binding of M-CSF to c-Fms a of resulting in formation of a multimeric complex for cell motility and cytoskeletal reorganization. The complex and its assembly activation of the c-Fms the of specific tyrosine residues in the cytoplasmic domain of the and of Bone marrow-derived macrophages from of and for 2 in with in the of of to M-CSF S. K. A. J. Bone Miner. Res. PubMed Scopus Google Scholar). and from of by of with for in the of of and R. Zallone A. Ross F.P. Teitelbaum S.L. J. 2003; PubMed Scopus Google Scholar) and in the of 2 and M-CSF for and of and of OCs and on in with or in the of for to or for to with various or and with Epo or M-CSF cells in with and protein for on with 2 of Ab, followed by with protein cell on protein In in followed by to with in at to and with for and with by with an and in with that are by and and the The used c-Fms and Vav3 by with Vav3 from followed by with OCs expressing various in the of and M-CSF to with M-CSF or Epo with the on OCs, and with and in a for at in in and on a an with and and chemotactic migration expressing the different both of with the in and and with and M-CSF. with in and used for cell or migration cell spreading, cells and for in the or absence of cells in and with migration cells in with and to for a gradient of Epo or on the of the at in a and the from of a Rac expressing the with Epo for and (12Faccio R. Novack D.V. Zallone A. Ross F.P. Teitelbaum S.L. J. Cell Biol. 2003; 162: 499-509Crossref PubMed Scopus (149) Google Scholar). to a Rac followed by against M-CSF-dependent in of PI3K to c-Fms PI3K M-CSF-dependent cytoskeletal S. R. Eur. J. 2001; PubMed Scopus Google Scholar, A. Y. I. K. K. PubMed Scopus Google Scholar), the c-Fms tyrosine that the cytoskeletal reorganization in to M-CSF in OCs not been this we a the domain of the to the transmembrane and cytoplasmic domain of osteoclast and macrophage c-Fms S. S. Teitelbaum S.L. Ross F.P. 2002; PubMed Scopus Google Scholar). We mutated the seven known Tyr residues in the cytoplasmic domain of c-Fms to Phe and a at the of the to with receptor or with mutated forms of with Epo, and and the with followed by Using an that the cytoplasmic domain of c-Fms and both the and c-Fms we that of both S. S. Teitelbaum S.L. Ross F.P. 2002; PubMed Scopus Google Scholar) not in the Tyr to Phe mutation that binding that Tyr-721 a binding we an receptor in which Tyr-721 to a of the in which all seven Tyr residues been by Phe add-back recruited the of PI3K the Furthermore, add-back Tyr-721, but not mutated Tyr-721, tyrosine for with Epo with the that two specific of or cytoskeletal S. R. Eur. J. 2001; PubMed Scopus Google Scholar), that of PI3K to c-Fms in M-CSF-dependent cytoskeletal this we OCs from with the or the in the of and M-CSF and cells with Epo to cytoskeletal reorganization dependent on the to the of M-CSF on OCs of Epo of cells expressing rapid podosome from a to the cell to membrane cells to respond that c-Fms Tyr-721 recruited PI3K, that the to Epo in a from to OCs, macrophages expressing to Epo with rapid in actin leading to of at the cell periphery WT, mutation of the c-Fms PI3K binding site Tyr-721 to Phe in from by c-Fms Tyr-559 but not for with M-CSF or and to with to actin OCs mutated and with Epo for and in A. an of a specific of the cell macrophages expressing or or the and to Epo for and membrane M-CSF-induced and in OCs c-Fms is the known of actin in OCs A. Y. I. K. K. PubMed Scopus Google Scholar, T. A. S. R. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, M. C. K. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). of c-Fms in that bind to c-Fms via Tyr-559 G. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). we that this Tyr important in the M-CSF-dependent of the actin in We that Tyr-559 an binding site in macrophages we cytoskeletal reorganization in OCs and expressing and that in both cell the mutated receptor to cytoskeletal reorganization in to Epo and specific for mutation in Tyr-559 all Tyr to Phe mutated forms of to actin remodeling M-CSF of cells to an PI3K J. Bone Miner. Res. PubMed Scopus Google Scholar, A. Y. I. K. K. PubMed Scopus Google Scholar, K. Jagger A. R. Chambers J. PubMed Scopus Google Scholar) and M. S. M. R. J. PubMed Scopus Google Scholar, T. A. S. R. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). we determined movement of cells of the via c-Fms Tyr-721 We the of OC expressing or mutated Tyr-721 and Tyr-559 by on and the of cells migrating or one the actin cell motility in cells and the of the PI3K and J. Bone Miner. Res. PubMed Scopus Google In contrast to the chemotactic Epo for the that binding of to c-Fms Tyr-559 for cytoskeletal and motility by the fact that the Src kinase migration of OC expressing receptor with the fact that inhibition of PI3K activation by also cell motility and membrane that both and PI3K M-CSF-dependent the binding of PI3K to c-Fms Tyr-721 not for M-CSF-induced cell motility and actin c-Fms Tyr-559 for cytoskeletal that PI3K cytoskeletal reorganization by binding to cytoplasmic Tyr residues in the c-Fms Tyr-721 the binding PI3K in to M-CSF this kinase recruited to the RTK by a complex with signaling molecules in M-CSF that and PI3K a signaling downstream of an with and PI3K in a c-Fms Tyr-559 is a known binding site for in macrophages G. M. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) and in the myeloid cell V. Cell. Biol. Res. 1999; PubMed Scopus Google Scholar). of macrophages and OCs on an and at T. A. S. R. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar), an that for of the of PI3K to the F. C.A. J. PubMed Scopus Google Scholar). we determined an complex recruited to c-Fms in OC by cells with the cell with and for in binding of to the c-Fms receptor upon M-CSF this in cells that binds Tyr-559 in c-Fms and that the and are known to in T. T. S. G. B. H. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar), we used receptor approach to the of Tyr-559 in of to the with and expressing with Epo and for binding of and of Epo and recruited to the the mutated of the same to WT, mutated receptor of the of and to c-Fms receptor Epo with the of a M.H. I. Nat. Cell. Biol. 2005; PubMed Scopus Google Scholar), the and within of Epo with in the mutated receptor The fact that Tyr-559 is a site for of an with the that has been to bind PI3K J. Biol. 1996; Full Text Full Text PDF PubMed Scopus Google Scholar), that formation of a complex and In with this and that binding of to in OC with with Epo, the same not in cells expressing c-Fms and not and binding of to is by the and in cells from Furthermore, of PI3K with of and is dependent on Tyr-559 that an the and PI3K, we used cells with M-CSF with and the and binding of and PI3K M-CSF and for at this in cells that members of the family bind to c-Fms Tyr-559 and recruit is to recruit PI3K to the Tyr-559 to that mutation of c-Fms Tyr-559 M-CSF-dependent cytoskeletal reorganization and binding of to and PI3K, we OC expressing an receptor in which Tyr-559 the tyrosine of c-Fms signals M-CSF-induced motility and cells expressing WT, or Tyr-559 add-back on in the of Epo to or in a in an Epo gradient to migration and membrane with membrane migration and the Tyr-559 add-back also of cell spreading, and membrane M-CSF-dependent cytoskeletal in macrophages V. H. Cell. Biol. 2005; PubMed Scopus (70) Google Scholar) and OCs R. Teitelbaum S.L. K. Zallone A. Tybulewicz Ross F.P. Nat. 2005; PubMed Scopus Google Scholar) via activation of Vav3 and its downstream Rac, we that this signaling dependent on In with this we that mutation of c-Fms Tyr-559 to Phe Vav3 and Rac the mutated receptor not the activation of two molecules Furthermore, reflecting its to cytoskeletal add-back Tyr-559 not sufficient to Vav3 and Rac activation In that Tyr-559 for M-CSF-dependent cytoskeletal reorganization in and OCs, this to cytoskeletal reorganization. Tyr-559 of an in the of c-Fms Tyr-697 and the of c-Fms Tyr residues Tyr-559 in M-CSF-dependent of the we add-back where Tyr-697 and Tyr-721 with the of in Tyr-697 to a binding site for P. T. J. PubMed Scopus Google Scholar). Tyr-697 to the cytoskeletal reorganization by M-CSF in cells the R. Zallone A. Ross F.P. Teitelbaum S.L. J. 2003; PubMed Scopus Google Scholar). Tyr-721 for its to a binding site for the of the we determined their to cell migration in to OC expressing WT, add-back or add-back and to a gradient of Tyr-559 add-back not OC migration in to Epo and the process by of or add-back migration The fact that Tyr-559 for binding and to this complex Tyr-559 or the Tyr residues of OC expressing receptor or the add-back Tyr-559 with Epo and by with a followed by for the of Tyr-559 mutated not induce binding of to its to cell the add-back receptor the complex to In that Tyr-559 a for SFK·c-Cbl·PI3K the formation of this complex the of Tyr-697 and In this we detailed the by which c-Fms M-CSF-dependent cell migration and cytoskeletal reorganization in macrophages and of PI3K to c-Fms Tyr-721, in the of a c-Fms receptor where all the Tyr residues are is not for M-CSF-induced cytoskeletal Tyr to Phe mutation of c-Fms Tyr-559 reorganization and OC migration in to M-CSF via inhibiting the formation of an SFK·c-Cbl·PI3K signaling complex and the activation of Vav3 and is not sufficient to M-CSF signaling to the a of Tyr-559 and Tyr-697 Tyr-721, with all Tyr residues mutated to can the formation of an complex and M-CSF-dependent both and actin remodeling and cell motility activation of specific receptor tyrosine Cell Res. 2002; PubMed Scopus Google Scholar, J. H. Curr. 2005; PubMed Scopus (70) Google Scholar). Cell migration and cytoskeletal reorganization are to macrophages and OCs activation of the M-CSF receptor Macrophages recruited to sites in to M-CSF and in the of their actin cytoskeleton. is in OCs migrating on bone in to a gradient of the same Bone a of initiated by binding of the cell to the via the and formation of a that the to where and are followed by and migration to a site on bone in to where the of is repeated I. J. 2003; PubMed Scopus Google Scholar, O. Saltel F. Jurdic P. F. Biol. Cell. 2003; 14: PubMed Scopus Google Scholar). The signals in actin can initiated by to the the and Src homology 2 domain-containing PI3K, and T. P. Trends Cell Biol. 2001; Full Text Full Text PDF PubMed Scopus Google Scholar). are recruited to a specific phospho-Tyr of to cell migration by cell or M. A. 2004; PubMed Scopus Google Scholar, Parsons PubMed Google Scholar). to PI3K, and of by of a phospho-Tyr within the of that for the Src homology 2 domain in of the T. Cell. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). the of c-Fms tyrosine residues in cell and signaling determined by wild-type or mutated c-Fms into or myeloid cells C.M. J. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar, K. A.R. Cell Google Scholar, D.J. Cell. Biol. PubMed Scopus Google Scholar). the receptor and cell c-Fms or can into OCs, the signaling by which M-CSF the receptor and its downstream Furthermore, the cell used in not the actin of the In contrast to all used OC which of c-Fms and respond to M-CSF by their actin cytoskeleton. the of cytoplasmic Tyr residues in we a receptor consisting of the domain of the to the transmembrane and cytoplasmic of at in this receptor to Epo by OCs with the S. S. Teitelbaum S.L. Ross F.P. 2002; PubMed Scopus Google Scholar). Epo of macrophages induces autophosphorylation of the Tyr residues within the c-Fms tail and activation of the and with and with the by M-CSF binding its receptor The S. R. J. Teitelbaum S.L. Ross F.P. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) and S. S. Teitelbaum S.L. Ross F.P. 2002; PubMed Scopus Google Scholar) in to M-CSF or Epo of macrophages that c-Fms and the c-Fms receptor the of model to M-CSF-dependent signaling in that the receptor is not OCs in the of M-CSF and Epo of macrophages or OCs with or Tyr within the cytoplasmic domain of of M-CSF-dependent We find that the c-Fms which not the to recruit in to M-CSF M-CSF-dependent membrane and cell is in with the of in actin in OCs M. S. M. R. J. PubMed Scopus Google Scholar, T. A. S. R. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). the known of PI3K in M-CSF-dependent signaling A. Y. I. K. K. PubMed Scopus Google Scholar), we find that c-Fms Tyr-721, the binding site for PI3K, is not for podosome or for the chemotactic to M-CSF. The fact that a specific PI3K blocks M-CSF-induced migration the c-Fms Tyr-721 to Phe mutation to the suggests that binding of PI3K to c-Fms is not for signaling to the cytoskeleton. mutation of c-Fms Tyr-559 blocks all M-CSF-dependent actin we that PI3K recruited to a signaling complex at the Tyr-559 recruited to not with PI3K this in to M-CSF not in that Tyr-559 is the for of cell motility and actin in macrophages and OCs by the formation of a complex and and and is a known of podosome in OCs and membrane in macrophages A. A. P. R. Biol. Cell. 2005; PubMed Scopus Google Scholar, A. A. E. C. E. J. J. R. J. Cell Biol. 2001; PubMed Scopus Google Scholar, E. S. F.J. Stanley E.R. C.A. G. J. Cell. 2003; PubMed Scopus Google Scholar) and for migration in both cell E. S. F.J. Stanley E.R. C.A. G. J. Cell. 2003; PubMed Scopus Google Scholar, R. A. K. H. R. Biol. 2003; PubMed Scopus Google Scholar). In the we the M-CSF signaling leading to cell motility and the of in to M-CSF different of the c-Fms We find that Tyr-559 the binding of to and PI3K and the to In with a that in OC binds via its Src homology domain in to leading to of T. A. S. R. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar), we find that M-CSF-induced and is dependent on Src kinase activity and is in cells expressing the c-Fms Furthermore, we that is for the M-CSF-dependent of and PI3K, this complex fails to in of the OC and macrophage on the activation of the small Rac, and which lead to actin polymerization through syndrome protein and the complex (11Calle Y. Jones G.E. Jagger C. Fuller K. Blundell M.P. Chow J. Chambers T. Thrasher A.J. Blood. 2004; 103: 3552-3561Crossref PubMed Scopus (98) Google Scholar, 14Hurst I.R. Zuo J. Jiang J. Holliday L.S. J. Bone Miner. Res. 2004; 19: 499-506Crossref PubMed Scopus (69) Google Scholar). M-CSF is a activator of Rac in OCs (12Faccio R. Novack D.V. Zallone A. Ross F.P. Teitelbaum S.L. J. Cell Biol. 2003; 162: 499-509Crossref PubMed Scopus (149) Google Scholar), a process dependent on the Vav3 R. Teitelbaum S.L. K. Zallone A. Tybulewicz Ross F.P. Nat. 2005; PubMed Scopus Google Scholar). of syndrome protein (11Calle Y. Jones G.E. Jagger C. Fuller K. Blundell M.P. Chow J. Chambers T. Thrasher A.J. Blood. 2004; 103: 3552-3561Crossref PubMed Scopus (98) Google Scholar) or Vav3 R. Teitelbaum S.L. K. Zallone A. Tybulewicz Ross F.P. Nat. 2005; PubMed Scopus Google Scholar), small interfering of complex I.R. Zuo J. Jiang J. Holliday L.S. J. Bone Miner. Res. 2004; 19: 499-506Crossref PubMed Scopus (69) Google Scholar), inhibition of Rac (13Razzouk S. Lieberherr M. Cournot G. Eur. J. Cell Biol. 1999; 78: 249-255Crossref PubMed Scopus (70) Google Scholar), or of M.A. S. S. Wang J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar) lead to OC and in macrophage that dynamic of the in cells is a for their function. We find that cells expressing c-Fms but not Vav3 and Rac that the SFK·c-Cbl·PI3K complex signals through that Tyr-559 is the site for the of cytoskeletal signaling in M-CSF-dependent of actin a c-Fms cytoplasmic domain in which Tyr-559 is the Tyr with all mutated to Phe induce actin reorganization and motility in to In contrast to we find that Tyr-559 is but not sufficient to initiate In Tyr-559 is the in the c-Fms the receptor is not of activation of Vav3 and Rac or the formation of the cell spreading, and We can the that the receptor this Tyr is not S. R. J. Teitelbaum S.L. Ross F.P. J. Biol. Full Text Full Text PDF PubMed Scopus Google Scholar). The fact that the SFK·c-Cbl·PI3K which fails to in cells expressing is also in cells expressing the Tyr-559 add-back suggests that Tyr residues are to or the Tyr-697 is a known binding site for and Tyr-721 that the of Tyr-697 and Tyr-721, in with cell motility to and formation of the complex and a model in which formation of a signaling complex of remodeling of the actin Tyr residues in c-Fms, and Tyr-559 the binding site for which are and and which in The fact that the complex SFK·c-Cbl·PI3K not in cells expressing a c-Fms receptor with the Tyr-559 suggests that the receptor signaling complex is by binding to Tyr-697 and of M-CSF-induced signaling to a that in cytoskeletal a key in OC and macrophage function. for this signaling is by the fact that of the receptor signaling complex is initiated by to the various residues Y. S. R. J. J. Biol. 2004; Full Text Full Text PDF PubMed Scopus Google Scholar). are to important for a process by reorganization of the actin cytoskeleton. In this the of various c-Fms tyrosine residues in the of M-CSF cytoskeletal and the mechanisms by which and PI3K are and recruited to specific c-Fms residues in a in the We for the

Source: Journal of Biological Chemistry

DOI: 10.1074/jbc.m610937200