Skip to main content

Advertisement

SpringerLink
Log in

The Signaling Adapter, FRS2, Facilitates Neuronal Branching in Primary Cortical Neurons via Both Grb2- and Shp2-Dependent Mechanisms

  • Published:
Journal of Molecular Neuroscience Aims and scope Submit manuscript

Abstract

The neurotrophins are a family of closely related growth factors that regulate proliferation and differentiation in the developing and mature nervous systems. Neurotrophins stimulate a family of receptor tyrosine kinases (Trk receptors) and utilize an intracellular docking protein termed fibroblast growth factor (FGF) receptor substrate 2 (FRS2) as a major downstream adapter to activate Ras, phosphatidylinositide 3-kinase (PI3K), and mitogen-activated protein kinase (MAPK) signaling cascades. The goals of this study were twofold: first, to investigate the complexity of neurotrophin-induced FRS2 interactions in primary cortical neurons and to determine which pathway(s) are important in regulating neuronal growth and, second, to determine whether the related signaling adapter, FRS3, stimulates neuron growth comparable to FRS2. We find that neurotrophin treatment of primary cortical neurons stimulates the tyrosine phosphorylation of FRS2 and the subsequent recruitment of Shp2, Grb2, and Gab2. With FRS2 mutants deficient in Grb2 or Shp2 binding, we demonstrate that FRS2 binds Gab1 and Gab2 through Grb2, providing an alternative route to activate PI3 kinase and Shp2. Using recombinant adenoviruses expressing FRS2, we demonstrate that FRS2 overexpression promotes neurite outgrowth and branching in cortical neurons relative to controls. In contrast, overexpression of FRS3 does not stimulate neuronal growth. Moreover, we find that while loss of Shp2, but not Grb2, reduces brain-derived neurotrophic factor (BDNF)-induced MAPK activation, the loss of either pathway impairs neuronal growth. Collectively, these experiments demonstrate that FRS2 functions as an adapter of a multiprotein complex that is activated by the Trk receptors and that the activation of both Grb2- and Shp2-dependent pathways facilitates cortical neuronal growth.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  • Alberti L, Borrello MG, Ghizzoni S, Torriti F, Rizzetti MG, Pierotti MA (1998) Grb2 binding to the different isoforms of Ret tyrosine kinase. Oncogene 17:1079–1087

    Article  CAS  PubMed  Google Scholar 

  • Atwal JK, Singh KK, Tessier-Lavigne M, Miller FD, Kaplan DR (2003) Semaphorin 3F antagonizes neurotrophin-induced phosphatidylinositol 3-kinase and mitogen-activated protein kinase kinase signaling: a mechanism for growth cone collapse. J Neurosci 23:7602–7609

    CAS  PubMed  Google Scholar 

  • Burgar HR, Burns HD, Elsden JL, Lalioti MD, Heath JK (2002) Association of the signaling adaptor FRS2 with fibroblast growth factor receptor 1 (Fgfr1) is mediated by alternative splicing of the juxtamembrane domain. J Biol Chem 277:4018–4023

    Article  CAS  PubMed  Google Scholar 

  • Cross MJ, Lu L, Magnusson P, Nyqvist D, Holmqvist K, Welsh M, Claesson-Welsh L (2002) The Shb adaptor protein binds to tyrosine 766 in the FGFR-1 and regulates the Ras/MEK/MAPK pathway via FRS2 phosphorylation in endothelial cells. Mol Biol Cell 13:2881–2893

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Dhalluin C, Yan SK, Plotnikova O, Lee KW, Zeng L, Kuti M, Mujtaba S, Goldfarb MP, Zhou MM (2000) Structural basis of SNT PTB domain interactions with distinct neurotrophic receptors. Mol Cell 6:921–929

    Article  CAS  PubMed  Google Scholar 

  • Dixon SJ, MacDonald JIS, Robinson KN, Kubu CJ, Meakin SO (2006) Trk receptor binding and neurotrophin/fibroblast growth factor (FGF)-dependent activation of the FGF receptor substrate (FRS)-3. B B A Mol Cell Res 1763:366–380

    CAS  Google Scholar 

  • Easton JB, Moody NM, Zhu X, Middlemas DS (1999) Brain-derived neurotrophic factor induces phosphorylation of fibroblast growth factor receptor substrate 2. J Biol Chem 274:11321–11327

    Article  CAS  PubMed  Google Scholar 

  • Easton JB, Royer AR, Middlemas DS (2006) The protein tyrosine phosphatase, Shp2, is required for the complete activation of the RAS/MAPK pathway by brain-derived neurotrophic factor. J Neurochem 97:834–845

    Article  CAS  PubMed  Google Scholar 

  • Evan GI, Lewis GK, Ramsay G, Bishop JM (1985) Isolation of monoclonal antibodies specific for human c-myc proto-oncogene product. Mol Cell Biol 5:3610–3616

    CAS  PubMed Central  PubMed  Google Scholar 

  • Gluzman Y (1981) SV40-transformed simian cells support the replication of early SV40 mutants. Cell 23:175–182

    Article  CAS  PubMed  Google Scholar 

  • Gorski JA, Zeiler SR, Tamowski S, Jones KR (2003) Brain-derived neurotrophic factor is required for the maintenance of cortical dendrites. J Neurosc 23:6856–6865

    CAS  Google Scholar 

  • Gotoh N, Manova K, Tanaka S, Murohashi M, Hadari Y, Lee A, Hanada Y, Hiroe T, Ito M, Kurihara T, Nakazato H, Shibuya M, Lax I, Lacy E, Schlessinger J (2005) The docking protein FRS2α is an essential component of multiple fibroblast growth factor responses during early mouse development. Mol Cell Biol 25:4105–4116

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Gu H, Neel BG (2003) The ‘Gab’ in signal transduction. Trends Cell Biol 13:122–130

    Article  CAS  PubMed  Google Scholar 

  • Hadari YR, Kouhara H, Lax I, Schlessinger J (1998) Binding of Shp2 tyrosine phosphatase to FRS-2 is essential for fibroblast growth factor-induced PC12 cell differentiation. Mol Cell Biol 18:3966–3973

    CAS  PubMed Central  PubMed  Google Scholar 

  • Hadari YR, Gotoh N, Kouhara H, Lax I, Schlessinger J (2001) Critical role for the docking-protein FRS2α in FGF receptor-mediated signal transduction pathways. Proc Natl Acad Sci U S A 98:8578–8583

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • He T-C, Zhou S, Costa LTD, Yu J, Kinzler KW, Vogelstein B (1998) A simplified system for generating recombinant adenoviruses. Proc Natl Acad Sci U S A 95:2509–2514

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Hempstead BL, Birge RB, Fajardo JE, Glassman R, Mahadeo D, Kraemer R, Hanafusa H (1994) Expression of the V-CRK oncogene product in PC12 cells results in rapid differentiation by both nerve growth factor and epidermal growth factor-dependent pathways. Mol Biol Cell 14:1964–1971

    CAS  Google Scholar 

  • Holgado-Madruga M, Moscatello DK, Emlet DR, Dieterich R, Wong AJ (1997) Grb2-associated binder-1 mediates phosphatidylinositol 3-kinase activation and the promotion of cell survival by nerve growth factor. Proc Natl Acad Sci U S A 94:12419–12424

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Hryciw T, MacDonald JIS, Phillips R, Seah C, Pasternak S, Meakin SO (2010) The fibroblast growth factor receptor substrate 3 adapater is a developmentally regulate microtubule-associated protein expressed in migrating and differentiated neurons. J Neurochem 112:924–939

    Article  CAS  PubMed  Google Scholar 

  • Kao S, Jaiswal RK, Kolch W, Landreth GE (2001) Identification of the mechanisms regulating the differential activation of the MAPK cascade by epidermal growth factor and nerve growth factor in PC12 cells. J Biol Chem 276:18169–18177

    Article  CAS  PubMed  Google Scholar 

  • Korhonen JM, Saïd FA, Wong AJ, Kaplan DR (1999) Gab1 mediates neurite outgrowth, DNA synthesis, and survival in PC12 cells. J Biol Chem 274:37307–37314

    Article  CAS  PubMed  Google Scholar 

  • Kouhara H, Hadari YR, Spivak-Kroizman T, Schilling J, Bar-Sagi D, Lax I, Schlessinger J (1997) A lipid-anchored Grb2-binding protein that links FGF-receptor activation to the Ras/MAPK signaling pathway. Cell 89:693–702

    Article  CAS  PubMed  Google Scholar 

  • Kurihara D, Yamashita T (2012) Chondroitin sulfate proteoglycans down-regulate spine formation in cortical neurons by targeting tropopmyosin-related kinase B (TrkB) protein. J Biol Chem 287:13822–13828

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Lax I, Wong A, Lamothe B, Lee A, Frost A, Hawes J, Schlessinger J (2002) The docking protein FRS2a controls a MAP kinase-mediated negative feedback mechanism for signaling by the FGF receptors. Mol Cell 10:709–719

    Article  CAS  PubMed  Google Scholar 

  • Liu H-Y, Meakin SO (2002) ShcB and ShcC activation by the Trk family of receptor tyrosine kinases. J Biol Chem 277:26046–26056

    Article  CAS  PubMed  Google Scholar 

  • Liu Y, Rohrschneider LR (2002) The gift of Gab. FEBS Lett 515:1–7

    Article  CAS  PubMed  Google Scholar 

  • Liu BA, Engelmann BW, Nash PD (2012) The language of SH2 domain interactions defines phosphotyrosine-mediated signal transduction. FEBS Lett 586:2597–2605

    Article  CAS  PubMed  Google Scholar 

  • McDougall K, Kubu C, Verdi JM, Meakin SO (2001) Developmental expression patterns of the signaling adapters FRS-2 and FRS-3 during early embryogenesis. Mech Dev 103:145–148

    Article  CAS  PubMed  Google Scholar 

  • Meakin SO, MacDonald JIS, Gryz EA, Kubu CJ, Verdi JM (1999) The signaling adapter protein FRS-2 competes with Shc for binding to TrkA: a model for discriminating proliferation and differentiation. J Biol Chem 274:9861–9870

    Article  CAS  PubMed  Google Scholar 

  • Ong SH, Guy GR, Hadair YR, Laks S, Gotoh N, Schlessinger J, Lax I (2000) FRS2 proteins recruit intracellular signaling pathways by binding to diverse targets of fibroblast growth factor and nerve growth factor receptors. Mol Cell Biol 20:979–989

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ong SH, Hadari YR, Gotoh N, Guy GR, Schlessinger J, Lax I (2001) Stimulation of phosphatidylinositol 3-kinase by fibroblast growth factor receptors is mediated by coordinated recruitment of multiple docking proteins. Proc Natl Acad Sci U S A 98:6074–6079

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Ridyard MS, Robbins SM (2003) Fibroblast growth factor-2-induced signaling through lipid raft-associated fibroblast growth factor receptor substrate 2 (FRS2). J Biol Chem 278:13803–13809

    Article  CAS  PubMed  Google Scholar 

  • Ringstedt T, Lagercrantz H, Persson H (1993) Expression of members of the trk family in the developing postnatal rat brain. Dev Brain Res 72:119–131

    Article  CAS  Google Scholar 

  • Simister PC, Feller SM (2012) Order and disorder in large multi-site docking proteins of the Gab family—implications for signalling complex formation and inhibitor design strategies. Mol Bio Syst 8:33–46

    CAS  Google Scholar 

  • Stoletov KV, Ratcliffe KE, Terman BI (2002) Fibroblast growth factor receptor substrate 2 participates in vascular endothelial growth factor-induced signaling. FASEB J 16:1283–1285

    CAS  PubMed  Google Scholar 

  • Wang X, Asmann YW, Erickson-Johnson MR, Oliveira JL, Zhang H, Moura RD, Lazar AJ, Lev D, Bill K, Lloyd RV, Yaszemski MJ, Maran A, Oliveira AM (2011) High-resolution genomic mapping reveals consistent amplification of the fibroblast growth factor receptor substrate 2 gene in well-differentiated and de-differentiated liposarcomas. Genes Chromosom Cancer 50:849–858

    Article  CAS  PubMed  Google Scholar 

  • Yamamoto S, Yoshino I, Shimazaki T, Murohashi M, Hevner RF, Lax I, Okano H, Shibuya M, Schlessinge J, Goth N (2005) Essential role of Shp2-binding sites on FRS2α for corticogenesis and for FGF2-dependent proliferation of neural progenitor cells. Proc Natl Acad Sci U S A 102:15983–15988

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Zeng G, Meakin SO (2002) Over-expression of the signaling adapter FRS-2 reconstitutes the cell cycle deficit of a nerve growth factor non-responsive TrkA receptor mutant. J Neurochem 81:820–831

    Article  CAS  PubMed  Google Scholar 

  • Zhang Y, McKeehan K, Lin Y, Zhang J, Wang F (2008) Fibroblast growth factor receptor 1 (FGFR1) tyrosine phosphorylation regulates binding of FGFR substrate 2α (FRS2α) but not FRS2β to the receptor. Mol Endocrinol 22:167–175

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  • Zhao C, Yu D-H, Shen R, Feng G-S (1999) Gab2, a new pleckstrin homology domain-containing adapter protein, acts to uncouple signaling from ERK kinase to Elk-1. J Biol Chem 274:19649–19654

    Article  CAS  PubMed  Google Scholar 

  • Zhou L, McDougall K, Kubu CJ, Verdi JM, Meakin SO (2003) Genomic organization and comparative sequence analysis of the mouse and human FRS2, FRS3 genes. Mol Biol Rep 30:15–25

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

This research was supported by an operating grant from the Canadian Institutes of Health Research (CIHR) to SOM.

Author information

Authors and Affiliations

  1. Laboratory of Neural Signaling, Molecular Medicine Research Group, The Robarts Research Institute, 1151 Richmond St. N, London, Ontario, N6A 5B7, Canada

    Li Zhou, Asghar Talebian & Susan O. Meakin

  2. The Department of Biochemistry, The University of Western Ontario, London, Ontario, N6A 5B7, Canada

    Asghar Talebian & Susan O. Meakin

  3. The Graduate Program in Neuroscience, The University of Western Ontario, London, Ontario, N6A 5B7, Canada

    Susan O. Meakin

Authors
  1. Li Zhou
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Asghar Talebian
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Susan O. Meakin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Susan O. Meakin.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Zhou, L., Talebian, A. & Meakin, S.O. The Signaling Adapter, FRS2, Facilitates Neuronal Branching in Primary Cortical Neurons via Both Grb2- and Shp2-Dependent Mechanisms. J Mol Neurosci 55, 663–677 (2015). https://doi.org/10.1007/s12031-014-0406-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12031-014-0406-4

Keywords

Search

Navigation