The Inositol 5′-Phosphatase SHIP-1 and the Src Kinase Lyn Negatively Regulate Macrophage Colony-stimulating Factor-induced Akt Activity

Upon encountering macrophage colony-stimulating factor (M-CSF), human monocytes undergo a series of cellular signaling events leading to an increase in Akt activity. However, the regulation of these events is not completely understood. Because the inositol 5′-phosphatase SHIP-1 is an important regulator of intracellular levels of phosphatidylinositol 3,4,5-trisphosphate, an important second messenger necessary for Akt activation, we hypothesized that SHIP-1 was involved in the regulation of M-CSF receptor (M-CSF-R)-induced Akt activation. In the human monocytic cell line, THP-1, SHIP-1 became tyrosine-phosphorylated following M-CSF activation in a Src family kinase-dependent manner. Transfection of 3T3-Fms cells, which express the human M-CSF-R, with wild-type SHIP-1 showed that SHIP-1 was necessary for the negative regulation of M-CSF-induced Akt activation. In THP-1 cells, SHIP-1 bound Lyn, independent of the kinase activity of Lyn, following M-CSF activation. Utilizing a glutathione S-transferase fusion protein, we found that SHIP-1 bound to Lyn via the SHIP-1 Src homology 2 domain. Furthermore, transfection of THP-1 cells with a wild-type SHIP-1 construct reduced NF-κB-dependent transcriptional activation of a reporter gene, whereas a SHIP-1 Src homology 2 domain construct resulted in an increase in NF-κB activation. Additionally, in 3T3-Fms cells, Lyn enhanced the ability of SHIP-1 to regulate Akt activation by stabilizing SHIP-1 at the cellular membrane. Finally, macrophages isolated from both SHIP-1- and Lyn-deficient mice exhibited enhanced Akt phosphorylation following M-CSF stimulation. These data provide the first evidence of the involvement of both SHIP-1 and Lyn in the negative regulation of M-CSF-R-induced Akt activation. Upon encountering macrophage colony-stimulating factor (M-CSF), human monocytes undergo a series of cellular signaling events leading to an increase in Akt activity. However, the regulation of these events is not completely understood. Because the inositol 5′-phosphatase SHIP-1 is an important regulator of intracellular levels of phosphatidylinositol 3,4,5-trisphosphate, an important second messenger necessary for Akt activation, we hypothesized that SHIP-1 was involved in the regulation of M-CSF receptor (M-CSF-R)-induced Akt activation. In the human monocytic cell line, THP-1, SHIP-1 became tyrosine-phosphorylated following M-CSF activation in a Src family kinase-dependent manner. Transfection of 3T3-Fms cells, which express the human M-CSF-R, with wild-type SHIP-1 showed that SHIP-1 was necessary for the negative regulation of M-CSF-induced Akt activation. In THP-1 cells, SHIP-1 bound Lyn, independent of the kinase activity of Lyn, following M-CSF activation. Utilizing a glutathione S-transferase fusion protein, we found that SHIP-1 bound to Lyn via the SHIP-1 Src homology 2 domain. Furthermore, transfection of THP-1 cells with a wild-type SHIP-1 construct reduced NF-κB-dependent transcriptional activation of a reporter gene, whereas a SHIP-1 Src homology 2 domain construct resulted in an increase in NF-κB activation. Additionally, in 3T3-Fms cells, Lyn enhanced the ability of SHIP-1 to regulate Akt activation by stabilizing SHIP-1 at the cellular membrane. Finally, macrophages isolated from both SHIP-1- and Lyn-deficient mice exhibited enhanced Akt phosphorylation following M-CSF stimulation. These data provide the first evidence of the involvement of both SHIP-1 and Lyn in the negative regulation of M-CSF-R-induced Akt activation. Human monocytes have an average life-span of 24–72 h (1Cline M.J. Lehrer R.I. Territo M.C. Golde D.W. Ann. Intern. Med. 1978; 88: 78-88Crossref PubMed Scopus (84) Google Scholar). In the absence of growth factors, such as M-CSF, 1The abbreviations used are: M-CSF, macrophage colony-stimulating factor; M-CSF-R, macrophage colony-stimulating factor receptor; SH, Src homology; PI, phosphatidylinositol; PI(4,5)P2, phosphatidylinositol 4,5-bisphosphate; PI(3,4,5)P3, phosphatidylinositol 3,4,5-trisphosphate; CML, chronic myelogenous leukemia; WT, wild-type; DMEM, Dulbecco's modified Eagle's medium; BMM, bone marrow-derived macrophage; TBS, Tris-buffered saline. monocytes undergo apoptosis (2Mangan D.F. Wahl S.M. J. Immunol. 1991; 147: 3408-3412PubMed Google Scholar, 3Mangan D.F. Welch G.R. Wahl S.M. J. Immunol. 1991; 146: 1541-1546PubMed Google Scholar, 4Marsh C.B. Pomerantz R.P. Parker J.M. Winnard A.V. Mazzaferri Jr., E.L. Moldovan N. Kelley T.W. Beck E. Wewers M.D. J. Immunol. 1999; 162: 6217-6225PubMed Google Scholar). As evidence of the important role of M-CSF in monocyte and macrophage homeostasis, M-CSF-deficient mice have deficiencies in both monocytes and macrophages (5Wiktor-Jedrzejczak W. Urbanowska E. Aukerman S.L. Pollard J.W. Stanley E.R. Ralph P. Ansari A.A. Sell K.W. Szperl M. Exp. Hematol. 1991; 19: 1049-1054PubMed Google Scholar). However, the process by which M-CSF induces monocyte survival and/or differentiation into macrophages and the signaling proteins that function as negative regulators of these events are not completely understood. The M-CSF receptor (M-CSF-R) is a receptor tyrosine kinase, in vivo expression of which is limited to monocytes, macrophages, proliferating smooth muscle cells, and cells of the female reproductive tract (6Inaba T. Yamada N. Gotoda T. Shimano H. Shimada M. Momomura K. Kadowaki T. Motoyoshi K. Tsukada T. Morisaki N. J. Biol. Chem. 1992; 267: 5693-5699Abstract Full Text PDF PubMed Google Scholar, 7Arceci R.J. Shanahan F. Stanley E.R. Pollard J.W. Proc. Natl. Acad. Sci. U. S. A. 1989; 86: 8818-8822Crossref PubMed Scopus (234) Google Scholar). Upon binding its ligand, M-CSF, the receptor dimerizes and undergoes auto- and trans-phosphorylation at specific tyrosine residues within the cytoplasmic domain of the receptor. These tyrosine residues, based upon the human M-CSF-R protein sequence, include tyrosine 561 (Tyr-561), Tyr-699, Tyr-708, Tyr-723, and Tyr-809, and bind specific signaling proteins once they become phosphorylated (8Alonso G. Koegl M. Mazurenko N. Courtneidge S.A. J. Biol. Chem. 1995; 270: 9840-9848Abstract Full Text Full Text PDF PubMed Scopus (177) Google Scholar, 9Courtneidge S.A. Dhand R. Pilat D. Twamley G.M. Waterfield M.D. Roussel M.F. EMBO J. 1993; 12: 943-950Crossref PubMed Scopus (204) Google Scholar, 10van der G.P. Hunter T. EMBO J. 1993; 12: 5161-5172Crossref PubMed Scopus (124) Google Scholar, 11van der G.P. Hunter T. Mol. Cell. Biol. 1990; 10: 2991-3002Crossref PubMed Scopus (43) Google Scholar, 12Reedijk M. 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We have previously shown that the M-CSF-R activates the serine/threonine kinase Akt in a PI 3-kinase-dependent manner to promote cellular survival (18Kelley T.W. Graham M.M. Doseff A.I. Pomerantz R.W. Lau S.M. Ostrowski M.C. Franke T.F. Marsh C.B. J. Biol. Chem. 1999; 274: 26393-26398Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar). The activation of PI 3-kinase converts phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2) to phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P3), leading to increased levels of the second messenger PI(3,4,5)P3. The SH2 domain-containing inositol 5′-phosphatase, SHIP-1, acts as a negative regulator of Akt phosphorylation by dephosphorylating PI(3,4,5)P3 to PI(3,4)P2 (19Damen J.E. Liu L. Rosten P. Humphries R.K. Jefferson A.B. Majerus P.W. Krystal G. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 1689-1693Crossref PubMed Scopus (566) Google Scholar, 20Lioubin M.N. Algate P.A. Tsai S. Carlberg K. Aebersold A. Rohrschneider L.R. Genes Dev. 1996; 10: 1084-1095Crossref PubMed Scopus (378) Google Scholar, 21Scharenberg A.M. El Hillal O. Fruman D.A. Beitz L.O. Li Z. Lin S. Gout I. Cantley L.C. Rawlings D.J. Kinet J.P. EMBO J. 1998; 17: 1961-1972Crossref PubMed Scopus (386) Google Scholar). The SHIP family of proteins contains two members: SHIP-1 and SHIP-2. SHIP-1 was first characterized as an ∼145-kDa protein that became tyrosine-phosphorylated upon interleukin-3 stimulation of a murine hematopoietic cell line (19Damen J.E. Liu L. Rosten P. Humphries R.K. Jefferson A.B. Majerus P.W. Krystal G. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 1689-1693Crossref PubMed Scopus (566) Google Scholar). Further studies showed that SHIP-1 has several alternatively spliced isoforms, including 135, 125, and 110 kDa (22Damen J.E. Liu L. Ware M.D. Ermolaeva M. Majerus P.W. Krystal G. Blood. 1998; 92: 1199-1205Crossref PubMed Google Scholar, 23Lucas D.M. Rohrschneider L.R. Blood. 1999; 93: 1922-1933Crossref PubMed Google Scholar). Initially, SHIP-1 was shown to be involved in the negative regulation of B-cell and IgE receptor signaling (24D'Ambrosio D. Fong D.C. Cambier J.C. Immunol. Lett. 1996; 54: 77-82Crossref PubMed Scopus (85) Google Scholar, 25Fong D.C. Malbec O. Arock M. Cambier J.C. Fridman W.H. Daeron M. Immunol. Lett. 1996; 54: 83-91Crossref PubMed Scopus (113) Google Scholar). A related gene product to SHIP-1 was also identified and was named SHIP-2 (26Pesesse X. Deleu S. De Smedt F. Drayer L. Erneux C. Biochem. Biophys. Res. Commun. 1997; 239: 697-700Crossref PubMed Scopus (199) Google Scholar). SHIP-2 is characterized by the same 5′-phosphatase activity as SHIP-1. However, whereas both SHIP-1 and SHIP-2 hydrolyze PI(3,4,5)P3 in vitro, only SHIP-1 hydrolyzes phosphatidylinositol D. A. S. H. S. P. Blood. 1999; 93: PubMed Google Scholar). Furthermore, SHIP-1, which is limited to cells of the hematopoietic SHIP-2 is in and muscle and the and from SHIP-1 in both the and (26Pesesse X. Deleu S. De Smedt F. Drayer L. Erneux C. Biochem. Biophys. Res. Commun. 1997; 239: 697-700Crossref PubMed Scopus (199) Google Scholar, T. J.A. J. Biol. Chem. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar). SHIP-1 has in and However, has the involvement of SHIP-1 in monocytes and the specific used by to regulate M-CSF-induced cellular the Src kinase Lyn has shown to SHIP-1 both in and in vivo H. A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (85) Google Scholar). However, the of phosphorylation is the tyrosine phosphorylation its activity its to the membrane. However, of is as the of SHIP-1, phosphatidylinositol 3,4,5-trisphosphate, is in the membrane. Because mice of cells and M-CSF is important in cell homeostasis, we hypothesized that SHIP-1 was involved in the regulation of M-CSF-induced cellular signaling in monocytes and to the of we that SHIP-1 tyrosine-phosphorylated upon M-CSF activation of THP-1 cells, and phosphorylation in a Src family kinase-dependent manner. Transfection of 3T3-Fms cells with SHIP-1 the of SHIP-1 in M-CSF-induced Akt activation. Additionally, following M-CSF stimulation of THP-1 cells, SHIP-1 with the Src kinase via the SH2 domain of SHIP-1, and was independent of the kinase activity of SHIP-1 reduced NF-κB-dependent transcriptional activation of a reporter gene in THP-1 Furthermore, in 3T3-Fms cells, Lyn enhanced the ability of SHIP-1 to regulate Akt activation by the of SHIP-1 to the membrane. Finally, macrophages isolated from both SHIP-1 Lyn-deficient mice exhibited enhanced Akt phosphorylation M-CSF stimulation in to macrophages isolated from wild-type These data provide the first evidence of the involvement of both Lyn and SHIP-1 in M-CSF-R leading to a SHIP-1 activation, and at the cell membrane. THP-1 cell line was from the in with and at cells a from Roussel and in with and human M-CSF was from The Src kinase and its from The SHIP-1 and Lyn used for from Additionally, a was from the Akt and Lyn from the Akt and from Cell and the SHIP-1 was from from and and bone marrow-derived macrophages from wild-type and mice J.E. Rosten P. R. P. S.M. A. F. Krystal G. Humphries R.K. Genes Dev. 1998; 12: PubMed Scopus Google and wild-type and Lyn-deficient mice cells with and in with and in a for with of M-CSF to the of the and to for h at with in Akt A and and cell the and used for M-CSF cells cells for h in with M-CSF and at in of and and and used for and cells at of a of was and at for two in of in of and for via to and in with for at in with at in and with of the at for in and by enhanced cells with M-CSF, in of and with of the SH2 domain and of a of at two with and in the same manner for as of an of to THP-1 cells in and at for to stimulation with reporter was from and was from SHIP-1 and the SHIP-1 SH2 domain into a from K. Lyn into was a from of The construct was a from R. Transfection of THP-1 and cells by with of wild-type SHIP-1 SHIP-1 SH2 of and 2 of to for transfection h and by M-CSF for h at The cells in of cell activity was the Transfection of to h to transfection in the of cells with of SHIP-1 and of for transfection of and for h in with from and for h to with the Lyn of SHIP-1, of Lyn, and of to cells and in the same manner as In cells SHIP-1 Lyn, was to the transfection to of 3T3-Fms cells with SHIP-1 Lyn as in of A 2 and and to at for to the at for and the was as the The was in A and in of for and at for The was as the SHIP-1 following M-CSF and Akt M-CSF-deficient mice have reduced of monocytes and macrophages and mice and of macrophages and cells into including the (5Wiktor-Jedrzejczak W. Urbanowska E. Aukerman S.L. Pollard J.W. Stanley E.R. Ralph P. Ansari A.A. Sell K.W. Szperl M. Exp. Hematol. 1991; 19: 1049-1054PubMed Google Scholar, J.E. Rosten P. R. P. S.M. A. F. Krystal G. Humphries R.K. Genes Dev. 1998; 12: PubMed Scopus Google we to the of SHIP-1 function following M-CSF stimulation. In the cell line, THP-1, which both SHIP-1 and the M-CSF-R, Akt phosphorylated following M-CSF activation an that is with Akt activity following growth factor stimulation D. L.R. T. C.B. A.B. F. 1997; PubMed Scopus Google Scholar, M. P. N. P. EMBO J. 1996; PubMed Scopus Google Scholar, A.B. C.B. P. Biol. 1997; Full Text Full Text PDF PubMed Google Scholar, A. Proc. Natl. Acad. Sci. U. S. A. 2001; PubMed Scopus Google Scholar). Additionally, SHIP-1 tyrosine-phosphorylated following M-CSF activation of THP-1 cells that THP-1 cells are a for M-CSF-R was to A and studies the of tyrosine phosphorylation SHIP-1 activity two H. A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (85) Google that not of SHIP-1 was the for of its activity. has shown that phosphorylation in SHIP-1 phosphorylation and activity. was in a by K.W. J. N. C. F. A.R. D.M. J. Exp. Med. PubMed Scopus Google Scholar, S. Kelley T. M. D. Mol. Cell. Biol. 1997; 17: PubMed Google which that SHIP-1 with in leading to the of signaling events. these events that signaling the M-CSF-R stimulates both signaling in of Akt activation, and negative signaling in of SHIP-1 Because Akt PI(3,4,5)P3 to to the and SHIP-1 the of PI(3,4,5)P3 by PI(3,4,5)P3 to SHIP-1 a role in Akt activation M-CSF activation. the of SHIP-1 in M-CSF-R-induced cellular we to 3T3-Fms These cells, which have in studies to M-CSF-R signaling (18Kelley T.W. Graham M.M. Doseff A.I. Pomerantz R.W. Lau S.M. Ostrowski M.C. Franke T.F. Marsh C.B. J. Biol. Chem. 1999; 274: 26393-26398Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar, M. D. Roussel M.F. J. Biol. Chem. 1999; 274: Full Text Full Text PDF PubMed Scopus Google are with the human M-CSF-R not SHIP-1, a for These cells with wild-type SHIP-1 and levels that transfection of 3T3-Fms cells with wild-type SHIP-1 and reduced levels following M-CSF activation in to cells that not with SHIP-1 and was for Akt to These data are and in in The the increase in following M-CSF of the and to In the absence of SHIP-1, the of M-CSF stimulation is In the of SHIP-1, the is only for that 3T3-Fms cells not express SHIP-1, we also of and cells and evidence of SHIP-1 expression only in the cells Furthermore, SHIP-1 became tyrosine-phosphorylated following M-CSF activation of 3T3-Fms cells not These data the of SHIP-1 in the negative regulation of M-CSF-induced Akt activation. The Src the of SHIP-1 following M-CSF of THP-1 SHIP-1 is phosphorylated following M-CSF activation of THP-1 and 3T3-Fms cells, we to the of studies of signaling the M-CSF-R have the involvement of the Src via tyrosine 561 S.A. Dhand R. Pilat D. Twamley G.M. Waterfield M.D. Roussel M.F. EMBO J. 1993; 12: 943-950Crossref PubMed Scopus (204) Google Scholar). H. A. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus (85) Google have shown that the Src kinase Lyn SHIP-1 both in and in Furthermore, Li E.L. M.J. E. Proc. Natl. Acad. Sci. U. S. A. 1999; PubMed Scopus Google have shown that Lyn-deficient of a negative role for Src kinase and a for the phosphorylation of SHIP-1. we the SHIP-1 was phosphorylated in a Src kinase-dependent manner. that of THP-1 cells with an of the Src to M-CSF activation reduced the phosphorylation of SHIP-1 with the of these is shown in and that of THP-1 cells with resulted in a in SHIP-1 phosphorylation of M-CSF with the these data not that Lyn is for the phosphorylation of SHIP-1, they that the Src are an important following M-CSF activation. SHIP-1 with Lyn M-CSF via the SH2 of of the data the of Lyn in both SHIP-1 and Akt activation, we to SHIP-1 with Lyn following M-CSF activation. THP-1 cells, we that SHIP-1 with Lyn following M-CSF activation The same was for SHIP-1 to Furthermore, was not by of THP-1 cells with the The same was also with SHIP-1 to These data that the of SHIP-1 and Lyn is independent of both the kinase activity of Lyn and the phosphorylation of SHIP-1. that the of SHIP-1 and Lyn was we to the SH2 domain of SHIP-1 was involved in binding we a fusion protein the SH2 domain of SHIP-1. We domain to THP-1 cell and found that the SH2 domain of SHIP-1 with Lyn following M-CSF activation of THP-1 cells with with SHIP-1, was not by of THP-1 cells with and the involvement of the SHIP-1 SH2 domain in M-CSF-R The SHIP-1 SH2 for NF-κB the of the SH2 domain of SHIP-1, we of a SHIP-1 SH2 domain We THP-1 cells with a construct with an of SHIP-1 the which with SHIP-1. We to the activity of SHIP-1 via the of NF-κB activity for two the transfection of THP-1 cells is and the is and has in studies the involvement of in NF-κB activation H. Marsh C.B. S. J. Biol. Chem. Full Text Full Text PDF PubMed Scopus Google Scholar, S. Marsh C.B. J. Immunol. PubMed Scopus Google Scholar). has shown that Akt is of NF-κB activation, SHIP-1 in a of D. A. Biol. 1999; Full Text Full Text PDF PubMed Scopus Google Scholar, J.A. 1999; PubMed Scopus Google Scholar, J.A. 1999; PubMed Scopus Google Scholar). that transfection of THP-1 cells with wild-type SHIP-1 reduced NF-κB transcriptional activity M-CSF activation. In transfection of the SH2 domain of SHIP-1 resulted in a increase in NF-κB transcriptional activity M-CSF activation with SHIP-1, that SHIP-1 is also important in the negative regulation of M-CSF-induced NF-κB activation, and that the SHIP-1 SH2 domain is an important of The Src Lyn the of SHIP-1 to by SHIP-1 at the The the of SHIP-1 in Akt levels NF-κB activation following M-CSF stimulation. We to the of Lyn in We the 3T3-Fms cells they not express SHIP-1 Transfection of these cells with Lyn with wild-type SHIP-1 and to the role of Lyn in the regulation of M-CSF-induced Akt activation. in the of Lyn, levels are reduced to a with cells Lyn 2 and These data are in In the absence of Lyn, levels are in cells with also for Lyn to that Lyn was into these cells the tyrosine phosphorylation of SHIP-1 in cells that with Lyn and was the phosphorylation of SHIP-1 in cells not with Lyn and The phosphorylation of SHIP-1 Lyn that in not with that SHIP-1 phosphorylated following M-CSF stimulation of 3T3-Fms cells to a in the in the of the of Lyn, we to Lyn has the of SHIP-1. We 3T3-Fms cells that with SHIP-1 and Lyn SHIP-1 and isolated the We SHIP-1 from the and the of SHIP-1 within the in the absence of In the of Lyn, the tyrosine phosphorylation of SHIP-1 following M-CSF activation was enhanced with the Lyn 2 and Furthermore, the of SHIP-1 within the was in the cells with Lyn that 3T3-Fms cells with SHIP-1 in the absence of These data that Lyn function to and SHIP-1 at the following M-CSF activation, a role for Src kinase in M-CSF-induced Akt activation. from SHIP-1- Lyn-deficient Akt we have of THP-1 cells 3T3-Fms cells to the of SHIP-1 and Lyn M-CSF-induced Akt activation. In we have characterized the of SHIP-1 and Lyn in THP-1 cells following M-CSF activation. we to the of SHIP-1 and Lyn in Akt activity. marrow-derived macrophages from and mice and with M-CSF for the The cells and the for via and for Akt from SHIP-1 wild-type mice showed a phosphorylation of Akt by a in phosphorylation However, macrophages isolated from exhibited a phosphorylation of in the absence of with M-CSF, and a enhanced phosphorylation of Akt M-CSF to The same for Akt to These data are in of Akt are as of to the Akt for both wild-type SHIP-1 and of M-CSF macrophages Akt with wild-type that cells not express SHIP-1. These data the of SHIP-1 in M-CSF-induced Akt activity. the of Lyn, we also isolated bone cells from Lyn-deficient and wild-type mice in the same manner as for the SHIP-1 isolated from wild-type mice a Akt by a isolated from Lyn-deficient mice also showed enhanced Akt levels M-CSF with wild-type to The same was for Akt to these data are in and are in the same manner as the data for the Lyn-deficient macrophages Akt of M-CSF with wild-type that Lyn-deficient cells not express These data provide evidence of the of Lyn in SHIP-1 function M-CSF activation and also of the of both SHIP-1 and Lyn in the negative regulation of M-CSF-induced Akt activity. The of was to the role of SHIP-1 in M-CSF-induced cell activation and to the by which SHIP-1 M-CSF-induced Akt activity. the monocytic cell line THP-1, we found that SHIP-1 bound the Src kinase Lyn following M-CSF stimulation via the SHIP-1 SH2 domain. the SHIP-1 and Lyn was M-CSF stimulation tyrosine phosphorylation with not a of to via M-CSF-R activation. Furthermore, the SHIP-1 and Lyn was independent of the Src kinase activity of Lyn, as the Src kinase not In reduced the tyrosine phosphorylation of SHIP-1, not only that Src kinase was also that the kinase activity of Lyn and the phosphorylation of SHIP-1 not necessary for Further studies to be to the tyrosine in Lyn that is involved in binding the SHIP-1 SH2 domain. is to that Lyn a to and SHIP-1 to the M-CSF stimulation to SHIP-1 to its Lyn-deficient macrophages exhibited a in SHIP-1 phosphorylation with the wild-type not Because SHIP-1 of a of important domain-containing including the serine/threonine kinase of SHIP-1 at the is The in SHIP-1 phosphorylation in the absence of Lyn from reduced of SHIP-1 at the following M-CSF stimulation. Furthermore, we also that M-CSF activation the phosphorylation of a specific tyrosine in Lyn, the with SHIP-1. Further studies are to these These data provide evidence of the role of Lyn as a negative regulator of M-CSF-induced cellular the of Lyn in both and negative regulation of signaling events the role of Src kinase in M-CSF-R signaling K.W. J. N. C. F. A.R. D.M. J. Exp. Med. PubMed Scopus Google Scholar, D.M. G.M. J. Exp. Med. 1998; PubMed Scopus Google Scholar, R.J. A.R. 1998; Full Text Full Text PDF PubMed Scopus (378) Google Scholar, F. P. 1997; Full Text Full Text PDF PubMed Scopus Google Scholar, Biol. 1998; Full Text Full Text PDF PubMed Scopus Google Scholar, H. K. I. T. J. Exp. Med. 1998; PubMed Scopus Google Scholar, R.J. Immunol. 1999; Google Scholar). these same J.W. P.W. J. Exp. Med. PubMed Scopus Google have shown that the Src kinase has a negative role in murine macrophages with to The evidence is the first of a negative function of the Src kinase Lyn in M-CSF-R The of SHIP-1 in cellular regulation and survival is in the These and of cells into including the and of J.E. Rosten P. R. P. S.M. A. F. Krystal G. Humphries R.K. Genes Dev. 1998; 12: PubMed Scopus Google Scholar). The cellular of to of that the absence of SHIP-1 in activation of Akt M-CSF stimulation and a role in the of macrophages in in with the the bone marrow-derived cells with M-CSF for and with with In to the wild-type macrophages from phosphorylation of Akt h of a of M-CSF stimulation from the SHIP-1 a role in the regulation of monocyte and macrophage survival in to M-CSF stimulation. Furthermore, macrophages from Lyn-deficient mice also enhanced phosphorylation of be in to SHIP-1, Lyn is also involved in the regulation of M-CSF-induced Akt activation. The to which Akt phosphorylation is enhanced in the Lyn-deficient macrophages is to the that is enhanced in the macrophages, the of Lyn in Furthermore, levels are reduced in such as in the of phosphatidylinositol 3,4,5-trisphosphate necessary for Akt activation. is also important to in chronic myelogenous that the of SHIP-1 to a role in cellular was in studies that a of cells, including with the and cells from with CML, SHIP-1 M. S. G. T. Algate P.A. Rohrschneider L.R. Mol. Cell. Biol. 1999; 19: PubMed Scopus Google Scholar). These cells also have activation of a for cellular survival a of regulation of PI(3,4,5)P3. Furthermore, in cell the of SHIP-1 to cellular the cells in are in these data a role for SHIP-1 in of cell In for the first that both SHIP-1 and Lyn regulate M-CSF-induced Akt activation. Further studies are to the role of the SHIP-1 with We and for and with the

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