Skip to main content

Advertisement

SpringerLink
Log in

Sprouty Genes Are Expressed in Osteoblasts and Inhibit Fibroblast Growth Factor-Mediated Osteoblast Responses

  • Published:
Calcified Tissue International Aims and scope Submit manuscript

Abstract

Fibroblast growth factors (FGFs) and fibroblast growth factor receptors (FGFRs) are major regulators of skeletal growth and development. Signal transduction via FGFRs is complex and mediates proliferation, differentiation, or migration depending upon the cellular context. Members of the Spry gene family antagonize the FGFR signal transduction pathway and inhibit lung morphogenesis, angiogenesis, and chondrogenesis. We examined the expression of Spry2 in the osteoblastic MC3T3-E1 cell line. MC3T3-E1 cells express Spry2 in response to FGF1 stimulation. Treatment of MC3T3-E1 cells with FGF1 results in the expression of Spry2 in a manner consistent with an early response gene. Pharmacological inhibitors of mitogen-activated protein kinase activation inhibit FGF1-induced expression of Spry2 mRNA. Transient overexpression of Spry2 in MC3T3-E1 resulted in decreased FGF1-mediated extracellular signal-regulated kinase phosphorylation and FGF1-stimulated osteopontin promoter activity. Furthermore, we show that Spry2 interacts with Raf-1 in a glutathione-S-transferase pulldown assay and that this interaction may involve multiple sites. Finally, Spry2 expression precedes the onset of the expression of osteoblast differentiation markers in an in vitro assay of primary osteoblast differentiation. Taken together, these results indicate that Spry2 expression is an early response to stimulation by FGF1 in MC3T3-E1 cells and acts as a feedback inhibitor of FGF1-induced osteoblast responses, possibly through interaction with Raf1.

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

Similar content being viewed by others

References

  1. Neilson KM, Friesel R (1996) Ligand-independent activation of fibroblast growth factor receptors by point mutations in the extracellular, transmembrane, and kinase domains. J Biol Chem 271:25049–25057

    CAS  PubMed  Google Scholar 

  2. Ornitz DM (2001) Regulation of chondrocyte growth and differentiation by fibroblast growth factor receptor 3. Novartis Found Symp 232:63–80, 272–282

    CAS  PubMed  Google Scholar 

  3. Naski MC, Ornitz DM (1998) FGF signaling in skeletal development. Front Biosci 3:D781–D794

    CAS  PubMed  Google Scholar 

  4. Colvin JS, Bohne BA, Harding GW, McEwen DG, Ornitz DM (1996) Skeletal overgrowth and deafness in mice lacking fibroblast growth factor receptor 3. Nat Genet 12:390–397

    Article  CAS  PubMed  Google Scholar 

  5. Rousseau F, el Ghouzzi V, Delezoide AL, Legeai-Mallet L, Le Merrer M, Munnich A, Bonaventure J (1996) Missense FGFR3 mutations create cysteine residues in thanatophoric dwarfism type I (TD1). Hum Mol Genet 5:509–512

    Article  CAS  PubMed  Google Scholar 

  6. Tavormina PL, Shiang R, Thompson LM, Zhu YZ, Wilkin DJ, Lachman RS, Wilcox WR, Rimoin DL, Cohn DH, Wasmuth JJ (1995) Thanatophoric dysplasia (types I and II) caused by distinct mutations in fibroblast growth factor receptor 3. Nat Genet 9:321–328

    Article  CAS  PubMed  Google Scholar 

  7. Tavormina PL, Bellus GA, Webster MK, Bamshad MJ, Fraley AE, McIntosh I, Szabo J, Jiang W, Jabs EW, Wilcox WR, Wasmuth JJ, Donoghue DJ, Thompson LM, Francomano CA (1999) A novel skeletal dysplasia with developmental delay and acanthosis nigricans is caused by a Lys650Met mutation in the fibroblast growth factor receptor 3 gene. Am J Hum Genet 64:722–731

    CAS  PubMed  Google Scholar 

  8. Bellus GA, Spector EB, Speiser PW, Weaver CA, Garber AT, Bryke CR, Israel J, Rosengren SS, Webster MK, Donoghue DJ, Francomano CA (2000) Distinct missense mutations of the FGFR3 lys650 codon modulate receptor kinase activation and the severity of the skeletal dysplasia phenotype. Am J Hum Genet 67:1411–1421

    Article  CAS  PubMed  Google Scholar 

  9. Muenke M, Schell U, Hehr A, et al. (1994) A common mutation in the fibroblast growth factor receptor 1 gene in Pfeiffer syndrome. Nat Genet 8:269–274

    Article  CAS  PubMed  Google Scholar 

  10. Meyers GA, Orlow SJ, Munro IR, Przylepa KA, Jabs EW (1995) Fibroblast growth factor receptor 3 (FGFR3) transmembrane mutation in Crouzon syndrome with acanthosis nigricans. Nat Genet 11:462–464

    Article  CAS  PubMed  Google Scholar 

  11. Bellus GA, Gaudenz K, Zackai EH, Clarke LA, Szabo J, Francomano CA, Muenke M (1996) Identical mutations in three different fibroblast growth factor receptor genes in autosomal dominant craniosynostosis syndromes. Nat Genet 14:174–176

    Article  CAS  PubMed  Google Scholar 

  12. Reardon W, Winter RM, Rutland P, Pulleyn LJ, Jones BM, Malcolm S (1994) Mutations in the fibroblast growth factor receptor 2 gene cause Crouzon syndrome. Nat Genet 8:98–103

    Article  CAS  PubMed  Google Scholar 

  13. Jabs EW, Li X, Scott AF, Meyers G, Chen W, Eccles M, Mao JI, Charnas LR, Jackson CE, Jaye M (1994) Jackson-Weiss and Crouzon syndromes are allelic with mutations in fibroblast growth factor receptor 2. Nat Genet 8:275–279

    Article  CAS  PubMed  Google Scholar 

  14. Wilkie AO, Slaney SF, Oldridge M, et al. (1995) Apert syndrome results from localized mutations of FGFR2 and is allelic with Crouzon syndrome. Nat Genet 9:165–172

    Article  CAS  PubMed  Google Scholar 

  15. Rutland P, Pulleyn LJ, Reardon W, et al. (1995) Identical mutations in the FGFR2 gene cause both Pfeiffer and Crouzon syndrome phenotypes. Nat Genet 9:173–176

    Article  CAS  PubMed  Google Scholar 

  16. Schell U, Hehr A, Feldman GJ, et al. (1995) Mutations in FGFR1 and FGFR2 cause familial and sporadic Pfeiffer syndrome. Hum Mol Genet 4:323–328

    CAS  PubMed  Google Scholar 

  17. Neilson KM, Friesel RE (1995) Constitutive activation of fibroblast growth factor receptor-2 by a point mutation associated with Crouzon syndrome. J Biol Chem 270:26037–26040

    Article  CAS  PubMed  Google Scholar 

  18. Naski MC, Wang Q, Xu J, Ornitz DM (1996) Graded activation of fibroblast growth factor receptor 3 by mutations causing achondroplasia and thanatophoric dysplasia. Nat Genet 13:233–237

    Article  CAS  PubMed  Google Scholar 

  19. Hacohen N, Kramer S, Sutherland D, Hiromi Y, Krasnow MA (1998) Sprouty encodes a novel antagonist of FGF signaling that patterns apical branching of the Drosophila airways. Cell 92:253–263

    Article  CAS  PubMed  Google Scholar 

  20. Warburton D, Schwarz M, Tefft D, Flores-Delgado G, Anderson KD, Cardoso WV (2000) The molecular basis of lung morphogenesis. Mech Dev 92:55–81

    Article  CAS  PubMed  Google Scholar 

  21. Placzek M, Skaer H (1999) Airway patterning: a paradigm for restricted signalling. Curr Biol 9:R506–R510

    Article  CAS  PubMed  Google Scholar 

  22. Reich A, Sapir A, Shilo B (1999) Sprouty is a general inhibitor of receptor tyrosine kinase signaling. Development 126:4139–4147

    CAS  PubMed  Google Scholar 

  23. Kramer S, Okabe M, Hacohen N, Krasnow MA, Hiromi Y (1999) Sprouty: a common antagonist of FGF and EGF signaling pathways in Drosophila. Development 126:2515–2525

    CAS  PubMed  Google Scholar 

  24. Minowada G, Jarvis LA, Chi CL, Neubuser A, Sun X, Hacohen N, Krasnow MA, Martin GR (1999) Vertebrate Sprouty genes are induced by FGF signaling and can cause chondrodysplasia when overexpressed. Development 126:4465–4475

    CAS  PubMed  Google Scholar 

  25. Tefft JD, Lee M, Smith S, Leinwand M, Zhao J, Bringas P Jr, Crowe DL, Warburton D (1999) Conserved function of mSpry-2, a murine homolog of Drosophila sprouty, which negatively modulates respiratory organogenesis. Curr Biol 9:219–222

    Article  CAS  PubMed  Google Scholar 

  26. Perl AK, Hokuto I, Impagnatiello MA, Christofori G, Whitsett JA (2003) Temporal effects of Sprouty on lung morphogenesis. Dev Biol 258:154–168

    Article  CAS  PubMed  Google Scholar 

  27. Wong ES, Fong CW, Lim J, Yusoff P, Low BC, Langdon WY, Guy GR (2002) Sprouty2 attenuates epidermal growth factor receptor ubiquitylation and endocytosis, and consequently enhances Ras/ERK signalling. EMBO J 21:4796–4808

    Article  CAS  PubMed  Google Scholar 

  28. Suzuki A, Guicheux J, Palmer G, Miura Y, Oiso Y, Bonjour JP, Caverzasio J (2002) Evidence for a role of p38 MAP kinase in expression of alkaline phosphatase during osteoblastic cell differentiation. Bone 30:91–98

    Article  CAS  PubMed  Google Scholar 

  29. Quarles LD, Yohay DA, Lever LW, Caton R, Wenstrup RJ (1992) Distinct proliferative and differentiated stages of murine MC3T3-E1 cells in culture: an in vitro model of osteoblast development. J Bone Miner Res 7:683–692

    CAS  PubMed  Google Scholar 

  30. Guo X, Zhang YP, Mitchell DA, Denhardt DT, Chambers AF (1995) Identification of a ras-activated enhancer in the mouse osteopontin promoter and its interaction with a putative ETS-related transcription factor whose activity correlates with the metastatic potential of the cell. Mol Cell Biol 15:476–487

    CAS  PubMed  Google Scholar 

  31. Kovalenko D, Yang X, Nadeau RJ, Harkins LK, Friesel R (2003) Sef inhibits fibroblast growth factor signaling by inhibiting FGFR1 tyrosine phosphorylation and subsequent ERK activation. J Biol Chem 278:14087–14091

    Article  CAS  PubMed  Google Scholar 

  32. Friesel R, Brown SA (1992) Spatially restricted expression of fibroblast growth factor receptor-2 during Xenopus development. Development 116:1051–1058

    CAS  PubMed  Google Scholar 

  33. Yang X, Kovalenko D, Nadeau RJ, Harkins LK, Mitchell J, Zubanova O, Chen PY, Friesel R (2004) Sef interacts with TAK1 and mediates JNK activation and apoptosis. J Biol Chem 279:38099–38102

    CAS  PubMed  Google Scholar 

  34. Ozaki K, Kadomoto R, Asato K, Tanimura S, Itoh N, Kohno M (2001) ERK pathway positively regulates the expression of Sprouty genes. Biochem Biophys Res Commun 285:1084–1088

    Article  CAS  PubMed  Google Scholar 

  35. Casci T, Vinos J, Freeman M (1999) Sprouty, an intracellular inhibitor of Ras signaling. Cell 96:655–665

    Article  CAS  PubMed  Google Scholar 

  36. Guy GR, Wong ES, Yusoff P, Chandramouli S, Lo TL, Lim J, Fong CW (2003) Sprouty: how does the branch manager work? J Cell Sci 116:3061–3068

    Article  CAS  PubMed  Google Scholar 

  37. Iwanami M, Hiromi Y, Okabe M (2005) Cell-type specific utilization of multiple negative feedback loops generates developmental constancy. Genes Cells 10:743–752

    Article  CAS  PubMed  Google Scholar 

  38. Mansukhani A, Bellosta P, Sahni M, Basilico C (2000) Signaling by fibroblast growth factors (FGF) and fibroblast growth factor receptor 2 (FGFR2)-activating mutations blocks mineralization and induces apoptosis in osteoblasts. J Cell Biol 149:1297–1308

    Article  CAS  PubMed  Google Scholar 

  39. Debiais F, Hott M, Graulet AM, Marie PJ (1998) The effects of fibroblast growth factor-2 on human neonatal calvaria osteoblastic cells are differentiation stage specific. J Bone Miner Res 13:645–654

    CAS  PubMed  Google Scholar 

  40. Delany AM, Canalis E (1998) Dual regulation of stromelysin-3 by fibroblast growth factor-2 in murine osteoblasts. J Biol Chem 273:16595–16600

    Article  CAS  PubMed  Google Scholar 

  41. Newberry EP, Willis D, Latifi T, Boudreaux JM, Towler DA (1997) Fibroblast growth factor receptor signaling activates the human interstitial collagenase promoter via the bipartite Ets-AP1 element. Mol Endocrinol 11:1129–1144

    Article  CAS  PubMed  Google Scholar 

  42. Strayhorn CL, Garrett JS, Dunn RL, Benedict JJ, Somerman MJ (1999) Growth factors regulate expression of osteoblast-associated genes. J Periodontol 70:1345–1354

    Article  CAS  PubMed  Google Scholar 

  43. Mason JM, Morrison DJ, Bassit B, Dimri M, Band H, Licht JD, Gross I (2004) Tyrosine phosphorylation of sprouty proteins regulates their ability to inhibit growth factor signaling: a dual feedback loop. Mol Biol Cell 15:2176–2188

    Article  CAS  PubMed  Google Scholar 

  44. Li X, Brunton VG, Burgar HR, Wheldon LM, Heath JK (2004) FRS2-dependent SRC activation is required for fibroblast growth factor receptor-induced phosphorylation of Sprouty and suppression of ERK activity. J Cell Sci 117:6007–6017

    CAS  PubMed  Google Scholar 

  45. Rubin C, Zwang Y, Vaisman N, Ron D, Yarden Y (2005) Phosphorylation of carboxyl-terminal tyrosines modulates the specificity of Sprouty-2 inhibition of different signaling pathways. J Biol Chem 280:9735–9744

    CAS  PubMed  Google Scholar 

  46. Li X, Wheldon L, Heath JK (2003) Sprouty: a controversial role in receptor tyrosine kinase signalling pathways. Biochem Soc Trans 31:1445–1446

    CAS  PubMed  Google Scholar 

  47. Kim HJ, Bar-Sagi D (2004) Modulation of signalling by Sprouty: a developing story. Nat Rev Mol Cell Biol 5:441–450

    Article  CAS  PubMed  Google Scholar 

  48. Fong CW, Leong HF, Wong ES, Lim J, Yusoff P, Guy GR (2003) Tyrosine phosphorylation of Sprouty2 enhances its interaction with c-Cbl and is crucial for its function. J Biol Chem 278:33456–33464

    Article  CAS  PubMed  Google Scholar 

  49. Hanafusa H, Torii S, Yasunaga T, Nishida E (2002) Sprouty1 and Sprouty2 provide a control mechanism for the Ras/MAPK signalling pathway. Nat Cell Biol 4:850–858

    Article  CAS  PubMed  Google Scholar 

  50. Yusoff P, Lao DH, Ong SH, Wong ES, Lim J, Lo TL, Leong HF, Fong CW, Guy GR (2002) Sprouty2 inhibits the Ras/MAP kinase pathway by inhibiting the activation of Raf. J Biol Chem 277:3195–3201

    Article  CAS  PubMed  Google Scholar 

  51. Sasaki A, Taketomi T, Kato R, Saeki K, Nonami A, Sasaki M, Kuriyama M, Saito N, Shibuya M, Yoshimura A (2003) Mammalian Sprouty4 suppresses Ras-independent ERK activation by binding to Raf1. Nat Cell Biol 5:427–432

    Article  CAS  PubMed  Google Scholar 

  52. Morriss-Kay GM, Iseki S, Johnson D (2001) Genetic control of the cell proliferation-differentiation balance in the developing skull vault: roles of fibroblast growth factor receptor signalling pathways. Novartis Found Symp 232:102–121

    CAS  PubMed  Google Scholar 

  53. Burke D, Wilkes D, Blundell TL, Malcolm S (1998) Fibroblast growth factor receptors: lessons from the genes. Trends Biochem Sci 23:59–62

    Article  CAS  PubMed  Google Scholar 

  54. Malcolm S, Reardon W (1996) Fibroblast growth factor receptor-2 mutations in craniosynostosis. Ann NY Acad Sci 785:164–170

    CAS  PubMed  Google Scholar 

  55. Lemonnier J, Hay E, Delannoy P, Fromigue O, Lomri A, Modrowski D, Marie PJ (2001) Increased osteoblast apoptosis in apert craniosynostosis: role of protein kinase C and interleukin-1. Am J Pathol 158:1833–1842

    CAS  PubMed  Google Scholar 

Download references

Acknowledgments

We thank Calvin Vary, Lucy Liaw, Volkhard Lindner, and Douglas Spicer for helpful comments throughout the course of this work. We thank Norma Albrecht for expert editorial assistance. This work was supported by NIH grant P20-15555 from the COBRE Program of the National Center for Research Resources and NIH grants DE13248 and HL65301 (to R.F.).

Author information

Author notes

  1. J. B. Webster

    Present address: Seattle Genetics, Inc., 21823 30th Drive SE, Bothell, WA, 98021, USA

Authors and Affiliations

  1. Center for Molecular Medicine, Maine Medical Center Research Institute, 81 Research Drive, Scarborough, ME, 04074, USA

    X. Yang, J. B. Webster, D. Kovalenko, R. J. Nadeau, O. Zubanova, P.-Y. Chen & R. Friesel

  2. Cooperative Graduate Program in Molecular Genetics and Cell Biology, University of Maine, Orono, ME, 04469, USA

    R. J. Nadeau, P.-Y. Chen & R. Friesel

Authors
  1. X. Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J. B. Webster
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. D. Kovalenko
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. R. J. Nadeau
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. O. Zubanova
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. P.-Y. Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. R. Friesel
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to R. Friesel.

Additional information

X. Yang, J. B. Webster and D. Kovaleno are contributed equally to this work.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yang, X., Webster, J.B., Kovalenko, D. et al. Sprouty Genes Are Expressed in Osteoblasts and Inhibit Fibroblast Growth Factor-Mediated Osteoblast Responses. Calcif Tissue Int 78, 233–240 (2006). https://doi.org/10.1007/s00223-005-0231-4

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00223-005-0231-4

Keywords

Search

Navigation