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PMID: 1372091 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

Interaction of phosphatidylinositol 3-kinase-associated p85 with epidermal growth factor and platelet-derived growth factor receptors.

Molecular and cellular biology ·Vol. 12 ·No. 3 ·1992-03-00 ·Pages 981-90

Hu P, Margolis B, Skolnik EY, Lammers R, Ullrich A, Schlessinger J

Abstract

One of the immediate cellular responses to stimulation by various growth factors is the activation of a phosphatidylinositol (PI) 3-kinase. We recently cloned the 85-kDa subunit of PI 3-kinase (p85) from a lambda gt11 expression library, using the tyrosine-phosphorylated carboxy terminus of the epidermal growth factor (EGF) receptor as a probe (E. Y. Skolnik, B. Margolis, M. Mohammadi, E. Lowenstein, R. Fischer, A. Drepps, A. Ullrich, and J. Schlessinger, Cell 65:83-90, 1991). In this study, we have examined the association of p85 with EGF and platelet-derived growth factor (PDGF) receptors and the tyrosine phosphorylation of p85 in 3T3 (HER14) cells in response to EGF and PDGF treatment. Treatment of cells with EGF or PDGF markedly increased the amount of p85 associated with EGF and PDGF receptors. Binding assays with glutathione S-transferase (GST) fusion proteins demonstrated that either Src homology region 2 (SH2) domain of p85 is sufficient for binding to EGF and PDGF receptors and that receptor tyrosine autophosphorylation is required for binding. Binding of a GST fusion protein expressing the N-terminal SH2 domain of p85 (GST-N-SH2) to EGF and PDGF receptors was half-maximally inhibited by 2 and 24 mM phosphotyrosine (P-Tyr), respectively, suggesting that the N-SH2 domain interacts more stably with PDGF receptors than with EGF receptors. The amount of receptor-p85 complex detected in HER14 cells treated with EGF or PDGF. Growth factor treatment also increased the amount of p85 found in anti-PDGF-treated HER14 cells, suggesting that the vast majority of p85 in the anti-P-Tyr fraction is receptor associated but not phosphorylated on tyrosine residues. Only upon transient overexpression of p85 and PDGF receptor did p85 become tyrosine phosphorylated. These are consistent with the hypothesis that p85 functions as an adaptor molecule that targets PI 3-kinase to activated growth factor receptors.

MeSH Terms
3T3 Cells Animals Cell Line ErbB Receptors/antagonists & inhibitors,metabolism Gene Expression Humans Immunoblotting Mice Phosphatidylinositol 3-Kinases Phosphotransferases/chemistry,genetics,metabolism Phosphotyrosine Platelet-Derived Growth Factor/metabolism Precipitin Tests Receptors, Cell Surface/antagonists & inhibitors,genetics,metabolism Receptors, Platelet-Derived Growth Factor Sulfhydryl Compounds/metabolism Tyrosine/analogs & derivatives,metabolism
Chemicals
Platelet-Derived Growth Factor Receptors, Cell Surface Sulfhydryl Compounds Phosphotyrosine Tyrosine Phosphotransferases Phosphatidylinositol 3-Kinases ErbB Receptors Receptors, Platelet-Derived Growth Factor
Authors & Affiliations
6 authors, click to expand affiliations / ORCID
Hu P
Department of Pharmacology, New York University Medical Center, New York 10016.
Margolis B
Skolnik E Y
Lammers R
Ullrich A
Schlessinger J
References (52)
52 references, click to expand
  1. Oncogenes and signal transduction.
    Cell. 1991 Jan 25;64(2):281-302 PMID: 1846320
  2. Evidence that the Rous sarcoma virus transforming gene product phosphorylates phosphatidylinositol and diacylglycerol.
    Proc Natl Acad Sci U S A. 1984 Apr;81(7):2117-21 PMID: 6326105
  3. Tyrosine kinase activity is essential for the association of phospholipase C-gamma with the epidermal growth factor receptor.
    Mol Cell Biol. 1990 Feb;10(2):435-41 PMID: 2153914
  4. Structural features of the colony-stimulating factor 1 receptor that affect its association with phosphatidylinositol 3-kinase.
    EMBO J. 1990 Aug;9(8):2415-21 PMID: 2164469
  5. Interactions of phosphatidylinositol kinase, GTPase-activating protein (GAP), and GAP-associated proteins with the colony-stimulating factor 1 receptor.
    Mol Cell Biol. 1990 Nov;10(11):5601-8 PMID: 2172781
  6. Purification and characterization of phosphoinositide 3-kinase from rat liver.
    J Biol Chem. 1990 Nov 15;265(32):19704-11 PMID: 2174051
  7. High-affinity epidermal growth factor binding is specifically reduced by a monoclonal antibody, and appears necessary for early responses.
    J Cell Biol. 1990 Feb;110(2):491-502 PMID: 2298813
  8. Common elements in growth factor stimulation and oncogenic transformation: 85 kd phosphoprotein and phosphatidylinositol kinase activity.
    Cell. 1987 Sep 25;50(7):1021-9 PMID: 2441878
  9. Role of phosphatidylinositol kinase in PDGF receptor signal transduction.
    Science. 1989 Mar 3;243(4895):1191-4 PMID: 2466336
  10. Phosphatidylinositol kinase or an associated protein is a substrate for the insulin receptor tyrosine kinase.
    J Biol Chem. 1990 Jan 5;265(1):396-400 PMID: 1688432
  11. A phosphatidylinositol-3 kinase binds to platelet-derived growth factor receptors through a specific receptor sequence containing phosphotyrosine.
    Mol Cell Biol. 1991 Feb;11(2):1125-32 PMID: 1703628
  12. Characterization of two 85 kd proteins that associate with receptor tyrosine kinases, middle-T/pp60c-src complexes, and PI3-kinase.
    Cell. 1991 Apr 5;65(1):91-104 PMID: 1707345
  13. EGF induces tyrosine phosphorylation of phospholipase C-II: a potential mechanism for EGF receptor signaling.
    Cell. 1989 Jun 30;57(7):1101-7 PMID: 2472218
  14. Phosphatidylinositol kinase type I activity associates with various oncogene products.
    Oncogene Res. 1989;4(4):283-92 PMID: 2549489
  15. Platelet-derived growth factor (PDGF) binding promotes physical association of PDGF receptor with phospholipase C.
    Proc Natl Acad Sci U S A. 1989 Nov;86(21):8232-6 PMID: 2554305
  16. Evidence for two distinct phosphatidylinositol kinases in fibroblasts. Implications for cellular regulation.
    Biochem J. 1987 Oct 1;247(1):165-74 PMID: 2825654
  17. Cloning and expression of a cDNA coding for the human platelet-derived growth factor receptor: evidence for more than one receptor class.
    Proc Natl Acad Sci U S A. 1988 May;85(10):3435-9 PMID: 2835772
  18. Cloning of bovine GAP and its interaction with oncogenic ras p21.
    Nature. 1988 Sep 1;335(6185):90-3 PMID: 2842690
  19. Association of phosphatidylinositol kinase activity with polyoma middle-T competent for transformation.
    Nature. 1985 May 16-22;315(6016):239-42 PMID: 2987699
  20. Non-catalytic domains of cytoplasmic protein-tyrosine kinases: regulatory elements in signal transduction.
    Oncogene. 1988 Nov;3(5):491-5 PMID: 3078956
  21. Point mutation at the ATP binding site of EGF receptor abolishes protein-tyrosine kinase activity and alters cellular routing.
    Cell. 1987 Oct 23;51(2):199-209 PMID: 3499230
  22. Transforming protein of avian sarcoma virus UR2 is associated with phosphatidylinositol kinase activity: possible role in tumorigenesis.
    Proc Natl Acad Sci U S A. 1984 May;81(9):2728-32 PMID: 6326140
  23. Increase of the catalytic activity of phospholipase C-gamma 1 by tyrosine phosphorylation.
    Science. 1990 Nov 30;250(4985):1253-6 PMID: 1700866
  24. The noncatalytic src homology region 2 segment of abl tyrosine kinase binds to tyrosine-phosphorylated cellular proteins with high affinity.
    Proc Natl Acad Sci U S A. 1991 Jan 15;88(2):627-31 PMID: 1703304
  25. Identification of domains of the v-crk oncogene product sufficient for association with phosphotyrosine-containing proteins.
    Mol Cell Biol. 1991 Mar;11(3):1607-13 PMID: 1705010
  26. PDGF stimulation of inositol phospholipid hydrolysis requires PLC-gamma 1 phosphorylation on tyrosine residues 783 and 1254.
    Cell. 1991 May 3;65(3):435-41 PMID: 1708307
  27. Activation of phosphatidylinositol 3-kinase in cells expressing abl oncogene variants.
    Mol Cell Biol. 1991 Feb;11(2):1107-13 PMID: 1846663
  28. cDNA cloning of a novel 85 kd protein that has SH2 domains and regulates binding of PI3-kinase to the PDGF beta-receptor.
    Cell. 1991 Apr 5;65(1):75-82 PMID: 1849460
  29. Nck, a melanoma cDNA encoding a cytoplasmic protein consisting of the src homology units SH2 and SH3.
    Nucleic Acids Res. 1990 Feb 25;18(4):1048 PMID: 2107526
  30. Activation of phosphatidylinositol 3-kinase by insulin.
    Proc Natl Acad Sci U S A. 1990 Feb;87(4):1411-5 PMID: 2154747
  31. Activated type I phosphatidylinositol kinase is associated with the epidermal growth factor (EGF) receptor following EGF stimulation.
    Proc Natl Acad Sci U S A. 1990 May;87(10):3816-20 PMID: 2160078
  32. Phosphorylation of the PDGF receptor beta subunit creates a tight binding site for phosphatidylinositol 3 kinase.
    EMBO J. 1990 Oct;9(10):3279-86 PMID: 2170111
  33. Binding of SH2 domains of phospholipase C gamma 1, GAP, and Src to activated growth factor receptors.
    Science. 1990 Nov 16;250(4983):979-82 PMID: 2173144
  34. The tyrosine phosphorylated carboxyterminus of the EGF receptor is a binding site for GAP and PLC-gamma.
    EMBO J. 1990 Dec;9(13):4375-80 PMID: 2176151
  35. Src homology region 2 domains direct protein-protein interactions in signal transduction.
    Proc Natl Acad Sci U S A. 1990 Nov;87(21):8622-6 PMID: 2236073
  36. Phosphatidylinositol metabolism and polyoma-mediated transformation.
    Proc Natl Acad Sci U S A. 1986 Jun;83(11):3624-8 PMID: 2424008
  37. An 81 kd protein complexed with middle T antigen and pp60c-src: a possible phosphatidylinositol kinase.
    Cell. 1987 Sep 25;50(7):1031-7 PMID: 2441879
  38. A novel viral oncogene with structural similarity to phospholipase C.
    Nature. 1988 Mar 17;332(6161):272-5 PMID: 2450282
  39. PDGF-dependent tyrosine phosphorylation stimulates production of novel polyphosphoinositides in intact cells.
    Cell. 1989 Apr 7;57(1):167-75 PMID: 2467744
  40. Phospholipase C-gamma is a substrate for the PDGF and EGF receptor protein-tyrosine kinases in vivo and in vitro.
    Cell. 1989 Jun 30;57(7):1109-22 PMID: 2472219
  41. Autophosphorylation of the PDGF receptor in the kinase insert region regulates interactions with cell proteins.
    Cell. 1989 Sep 22;58(6):1121-33 PMID: 2550144
  42. Platelet-derived growth factor induces rapid and sustained tyrosine phosphorylation of phospholipase C-gamma in quiescent BALB/c 3T3 cells.
    Mol Cell Biol. 1989 Jul;9(7):2934-43 PMID: 2550789
  43. The colony stimulating factor-1 receptor associates with and activates phosphatidylinositol-3 kinase.
    Nature. 1989 Dec 7;342(6250):699-702 PMID: 2556641
  44. Sequence similarity of phospholipase C with the non-catalytic region of src.
    Nature. 1988 Mar 17;332(6161):269-72 PMID: 2831461
  45. Type I phosphatidylinositol kinase makes a novel inositol phospholipid, phosphatidylinositol-3-phosphate.
    Nature. 1988 Apr 14;332(6165):644-6 PMID: 2833705
  46. Inositol phospholipid-specific phospholipase C: complete cDNA and protein sequences and sequence homology to tyrosine kinase-related oncogene products.
    Proc Natl Acad Sci U S A. 1988 Aug;85(15):5419-23 PMID: 2840660
  47. cDNA cloning and expression of a human platelet-derived growth factor (PDGF) receptor specific for B-chain-containing PDGF molecules.
    Mol Cell Biol. 1988 Aug;8(8):3476-86 PMID: 2850496
  48. A noncatalytic domain conserved among cytoplasmic protein-tyrosine kinases modifies the kinase function and transforming activity of Fujinami sarcoma virus P130gag-fps.
    Mol Cell Biol. 1986 Dec;6(12):4396-408 PMID: 3025655
  49. Single-step purification of polypeptides expressed in Escherichia coli as fusions with glutathione S-transferase.
    Gene. 1988 Jul 15;67(1):31-40 PMID: 3047011
  50. Molecular cloning of two types of GAP complementary DNA from human placenta.
    Science. 1988 Dec 23;242(4886):1697-700 PMID: 3201259
  51. High-efficiency transformation of mammalian cells by plasmid DNA.
    Mol Cell Biol. 1987 Aug;7(8):2745-52 PMID: 3670292
  52. Cloning of PI3 kinase-associated p85 utilizing a novel method for expression/cloning of target proteins for receptor tyrosine kinases.
    Cell. 1991 Apr 5;65(1):83-90 PMID: 1849461
Article Info
Journal
Molecular and cellular biology
Abbr.
Mol Cell Biol
ISSN
0270-7306
DOI
10.1128/mcb.12.3.981-990.1992
Published
1992-03-00
Pages
981-90
Language
English
Region
United States
NLM ID
8109087
PMCID
PMC369530
Subset
IM
Grants
NIDDK NIH HHS · DK01927 · United States
External Links
PubMed source DOI: 10.1128/mcb.12.3.981-990.1992 PMC Full Text View journal details
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