Novel Adapter Proteins that Link the Human GM-CSF Receptor to the Phosphatidylino-sitol 3-Kinase and Shc/Grb2/ras Signaling Pathways
We have used a human GM-CSF-dependent hematopoietic cell line that responds to physiological concentrations of hGM-CSF to analyze a set of signaling events that occur in normal myelopoiesis and whose deregulation may lead to leukemogenesis. Stimulation of these cells with hGM-CSF induced the assembly of multimeric complexes that contained known and novel phosphotyrosyl proteins. One of the new proteins was a major phosphotyrosyl substrate of 76–85 kDa (p80) that was directly associated with the p85 subunit of phosphatidylinositol (PI) 3-kinase through the SH2 domains of p85. p80 also associated with the β subunit of the activated hGM-CSF receptor, and assembly of this complex correlated with activation of PI 3-kinase. A second phosphotyrosyl protein we identified, pi40, associated with the She and Grb2 adapter proteins by direct binding to a novel phosphotyrosine-interacting domain located at the N-terminus of She, and to the SH3 domains of Grb2, respectively. The Shc/pl40/Grb2 complex was found to be constitutively activated in acute myeloid leukemia cells, indicating that activation of this pathway may be a necessary step in the development of some leukemias. The p80/p85/PI 3-kinase and the Shc/Grb2/pl40 complexes were tightly associated with Src family kinases, which were prime candidates for phosphorylation of She, p80, pl40 and other phosphotyrosyl substrates pesent in these complexes. Our studies suggest that p80 and pl40 may link the hGM-CSF receptor to the PI 3-kinase and Shc/Grb2/ras signaling pathways, respectively, and that abnormal activation of hGM-CSF-dependent targets may play a role in leukemogenesis.
KeywordsAgar Tyrosine Leukemia Glutathione Auger
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- 5.Corey S, Eguinoa A, Puyana-Theall K, Bolen JB, Cantley L, Molinedo F, Jackson TR, Hawkins PT, Stephens LR (1993) Granulocyte macrophage-colony stimulating factor stimulates both association and activation of phosphoinositide 30H-kinase and src-related tyrosine kinase(s) in human myeloid derived cells. EMBOJ 12:2681–2690Google Scholar
- 9.Hayashida K, Kitamura T, Gorman DM, Arai KI, Yokota T, Miyajima A (1990) Molecular cloning of a second subunit of the receptor for human granulocyte-macrophage colony stimulating factor (GM-CSF): reconstitution of a high-affinity GM-CSF receptor. Proc Natl Acad Sci USA 87:9655–9659PubMedCrossRefGoogle Scholar
- 11.Jücker M, Feldman RA, SubmittedGoogle Scholar
- 13.Kavanough WM, Klippel A, Escobedo JA, Williams LT (1992) Modification of the 85 kilodalton subunit of phosphatidylinositol-3 kinase in platelet-derived growth factor-stimulated cells. Mol Cell Biol 12:3415–3424Google Scholar
- 14.Keegan AD, Nelms K, White M,Wang L-M, Pierce JH, Paul,W (1994) An EL-4 receptor region containing an insulin receptor motif is important for IL-4-mediated IRS-1 phosphorylation and cell growth. Cell 76:811–820Google Scholar
- 18.Lanfrancone L, Pelicci G, Brizzi MF, Arouica MG, Casciari C, Giuli S, Pegoraro L, Pawson T, Pelicci PG (1995) Overexpression of She proteins potentiates the proliferative response to the granulocyte-macrophage colony-stimulating factor and recruitment of Grb2/SoS and Grb2/pl40 complexes to the receptor p subunit. Oncogene 10:907–917PubMedGoogle Scholar
- 19.Lowenstein EJ, Daly RJ, Batzer AG, Li W, Margolis B, LammersR, Ullrich A, Skolnik EY, Bar-Sagi D, Schlessinger J (1992) The SH2 and SH3 domain-containing protein GRB2 links receptor tyrosine kinases to ras signaling. Cell 70:431–442Google Scholar
- 26.Quelle FW, Shimoda K, Thierfelder W, Fischer, C, Kim A, Ruben SM, Cleveland JM, Pierce JH, Keegan AD, Nelms K, Paul WE, Ihle JN (1995) Cloning of murine Stat6 and human Stat6, Stat proteins mat are tyrosine phosphorylated in responses to DL-4 and EL-3 but are not required for mitogenesis. Mol Cell Biol 15:3336–3343PubMedGoogle Scholar
- 27.Rozakis-Adcock M, McGlade J, Mbamalu G, Pelicci G, Daly R., Li W, Batzer A, Thomas S, Brugge J, Pelicci P G, Schlessinger J, Pawson T (1992) Association of the She and Grb2/Sem5 SH2-containing proteins is implicated in activation of the ras pathway by tyrosine kinases. Nature 360:689–692PubMedCrossRefGoogle Scholar
- 30.Skolnik EY, Lee C-H, Batzer A, Vincentini LM, Zhou M, Daly R, Myers Jr MJ, Backer JM, Ullrich A, White MF, Schlessinger J (1993) The SH2/SH3 domain-containing protein GRB2 interacts with tyrosine-phosphorylated IRS 1 and She: implications for insulin control of ras signalling. EMBOJ 12:1929–1936Google Scholar