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Repelling class discrimination: ephrin-A5 binds to and activates EphB2 receptor signaling

Nature Neuroscience volume 7, pages 501–509 (2004)Cite this article

Abstract

The interactions between Eph receptor tyrosine kinases and their ephrin ligands regulate cell migration and axon pathfinding. The EphA receptors are generally thought to become activated by ephrin-A ligands, whereas the EphB receptors interact with ephrin-B ligands. Here we show that two of the most widely studied of these molecules, EphB2 and ephrin-A5, which have never been described to interact with each other, do in fact bind one another with high affinity. Exposure of EphB2-expressing cells to ephrin-A5 leads to receptor clustering, autophosphorylation and initiation of downstream signaling. Ephrin-A5 induces EphB2-mediated growth cone collapse and neurite retraction in a model system. We further show, using X-ray crystallography, that the ephrin-A5–EphB2 complex is a heterodimer and is architecturally distinct from the tetrameric EphB2–ephrin-B2 structure. The structural data reveal the molecular basis for EphB2–ephrin-A5 signaling and provide a framework for understanding the complexities of functional interactions and crosstalk between A- and B-subclass Eph receptors and ephrins.

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Figure 1: EphB2 and ephrin-A5 bind each other with high affinity.
Figure 2: Ephrin-A5 induces clustering and phosphorylation of the EphB2 receptor tyrosine kinase.
Figure 3: Ephrin-A5 induces neurite retraction in NG108-EphB2 cells.
Figure 4: Structure of the ephrin-A5–EphB2 complex.

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References

  1. Eph Nomenclature Committee. Unified nomenclature for Eph family receptors and their ligands, the Ephrins. Cell 90, 403–404 (1997).

  2. Flanagan, J.G. & Vanderhaeghen, P. The ephrins and Eph receptors in neural development. Annu. Rev. Neurosci. 21, 309–345 (1998).

    Article  CAS  Google Scholar 

  3. Frisen, J., Holmberg, J. & Barbacid, M. Ephrins and their Eph receptors: multitalented directors of embryonic development. EMBO J. 18, 5159–5165 (1999).

    Article  CAS  Google Scholar 

  4. Kullander, K. & Klein, R. Mechanisms and functions of Eph and ephrin signalling. Nat. Rev. Mol. Cell Biol. 3, 475–486 (2002).

    Article  CAS  Google Scholar 

  5. Henkemeyer, M. et al. Nuk controls pathfinding of commissural axons in the mammalian central nervous system. Cell 86, 35–46 (1996).

    Article  CAS  Google Scholar 

  6. Holland, S.J. et al. Bidirectional signalling through the EPH-family receptor Nuk and its transmembrane ligands. Nature 383, 722–725 (1996).

    Article  CAS  Google Scholar 

  7. Bruckner, K., Pasquale, E.B. & Klein, R. Tyrosine phosphorylation of transmembrane ligands for Eph receptors. Science 275, 1640–1643 (1997).

    Article  CAS  Google Scholar 

  8. Pasquale, E.B. The Eph family of receptors. Curr. Opin. Cell Biol. 9, 608–615 (1997).

    Article  CAS  Google Scholar 

  9. Gale, N.W. et al. Eph receptors and ligands comprise two major specificity subclasses and are reciprocally compartmentalized during embryogenesis. Neuron 17, 9–19 (1996).

    Article  CAS  Google Scholar 

  10. Gale, N.W. et al. Elk-L3, a novel transmembrane ligand for the Eph family of receptor tyrosine kinases, expressed in embryonic floor plate, roof plate and hindbrain segments. Oncogene 13, 1343–1352 (1996).

    CAS  PubMed  Google Scholar 

  11. Bergemann, A.D. et al. Ephrin-B3, a ligand for the receptor EphB3, expressed at the midline of the developing neural tube. Oncogene 16, 471–480 (1998).

    Article  CAS  Google Scholar 

  12. Beckmann, M.P. et al. Molecular characterization of a family of ligands for eph-related tyrosine kinase receptors. EMBO J. 13, 3757–3762 (1994).

    Article  CAS  Google Scholar 

  13. Kozlosky, C.J. et al. Ligands for the receptor tyrosine kinases hek and elk: isolation of cDNAs encoding a family of proteins. Oncogene 10, 299–306 (1995).

    CAS  PubMed  Google Scholar 

  14. Himanen, J.P., Henkemeyer, M. & Nikolov, D.B. Crystal structure of the ligand-binding domain of the receptor tyrosine kinase EphB2. Nature 396, 486–491 (1998).

    Article  CAS  Google Scholar 

  15. Toth, J. et al. Crystal structure of an ephrin ectodomain. Dev. Cell 1, 83–92 (2001).

    Article  CAS  Google Scholar 

  16. Himanen, J.P. et al. Crystal structure of an Eph receptor-ephrin complex. Nature 414, 933–938 (2001).

    Article  CAS  Google Scholar 

  17. Himanen, J.P. & Nikolov, D.B. Eph signaling: a structural view. Trends Neurosci. 26, 46–51 (2003).

    Article  CAS  Google Scholar 

  18. Mann, F., Miranda, E., Weinl, C., Harmer, E. & Holt, C.E. B-type Eph receptors and ephrins induce growth cone collapse through distinct intracellular pathways. J. Neurobiol. 57, 323–336 (2003).

    Article  CAS  Google Scholar 

  19. Marston, D.J., Dickinson, S. & Nobes, C.D. Rac dependent trans-endocytosis of ephrinBs regulates Eph-ephrin contact repulsion. Nat. Cell Biol. 5, 879–888 (2003).

    Article  CAS  Google Scholar 

  20. Zimmer, M., Palmer, A., Köhler, J. & Klein, R. EphB/ephrinB bi-directional endocytosis terminates adhesion allowing contact mediated repulsion. Nat. Cell Biol. 5, 869–878 (2003).

    Article  CAS  Google Scholar 

  21. Binns, K.L., Taylor, P.P., Sicheri, F., Pawson, T. & Holland, S.J. Phosphorylation of tyrosine residues in the kinase domain and juxtamembrane region regulates the biological and catalytic activities of Eph receptors. Mol. Cell. Biol. 20, 4791–4805 (2000).

    Article  CAS  Google Scholar 

  22. Holland, S. et al. Juxtamembrane tyrosine residues couple the Eph family receptor EphB2/Nuk to specific SH2 domain proteins in neuronal cells. EMBO J. 16, 3877–3888 (1997).

    Article  CAS  Google Scholar 

  23. Himanen, J.P. & Nikolov, D.B. Purification, crystallization and preliminary characterization of an Eph-B2/ephrin-B2 complex. Acta Crystallogr. D 58, 533–535 (2002).

    Article  Google Scholar 

  24. Koolpe, M., Dail, M. & Pasquale, E.B. An ephrin mimetic peptide that selectively targets the EphA2 receptor. J. Biol. Chem. 277, 46974–46979 (2002).

    Article  CAS  Google Scholar 

  25. Feldheim, D.A. et al. Genetic analysis of ephrin-A2 and ephrin-A5 shows their requirement in multiple aspects of retinocollicular mapping. Neuron 25, 563–574 (2000).

    Article  CAS  Google Scholar 

  26. Hindges, R., McLaughlin, T., Genoud, N., Henkemeyer, M. & O'Leary, D.D. EphB forward signaling controls directional branch extension and arborization required for dorsal-ventral retinotopic mapping. Neuron 35, 475–487 (2002).

    Article  CAS  Google Scholar 

  27. Knoll, B., Zarbalis, K., Wurst, W. & Drescher, U. A role for the EphA family in the topographic targeting of vomeronasal axons. Development 128, 895–906 (2001).

    CAS  PubMed  Google Scholar 

  28. Cutforth, T. et al. Axonal ephrin-As and odorant receptors. Coordinate determination of the olfactory sensory map. Cell 114, 311–322 (2003).

    Article  CAS  Google Scholar 

  29. St. John, J.A. & Key, B. EphB2 and two of its ligands have dynamic protein expression patterns in the developing olfactory system. Brain Res. Dev. Brain Res. 126, 43–56 (2001).

    Article  CAS  Google Scholar 

  30. Smith, F.M. et al. Dissecting the EphA3/ephrin-A5 interactions using a novel functional mutagenesis screen. J. Biol. Chem. 279, 9522–9531 (2004).

    Article  CAS  Google Scholar 

  31. Lackmann, M. et al. Distinct subdomains of the EphA3 receptor mediate ligand binding and receptor dimerization. J. Biol. Chem. 273, 20228–20237 (1998).

    Article  CAS  Google Scholar 

  32. Stapleton, D., Balan, I., Pawson, T. & Sicheri, F. The crystal structure of an Eph receptor SAM domain reveals a mechanism for modular dimerization. Nat. Struct. Biol. 6, 44–49 (1999).

    Article  CAS  Google Scholar 

  33. Thanos, C.D., Goodwill, K.E. & Bowie, J.U. Oligomeric structure of the human EphB2 receptor SAM domain. Science 283, 833–836 (1999).

    Article  CAS  Google Scholar 

  34. Lackmann, M. Isolation and characterization of “orphan-RTK” ligands using an integrated biosensor approach. Methods Mol. Biol. 124, 335–359 (2001).

    CAS  PubMed  Google Scholar 

  35. Okada, T. et al. Sindbis viral-mediated expression of Ca2+-permeable AMPA receptors at hippocampal CA1 synapses and induction of NMDA receptor-independent long-term potentiation. Eur. J. Neurosci. 13, 1635–1643 (2001).

    Article  CAS  Google Scholar 

  36. Cowan, C.A. & Henkemeyer, M. The SH2/SH3 adaptor Grb4 transduces B-ephrin reverse signals. Nature 413, 174–179 (2001).

    Article  CAS  Google Scholar 

  37. Henkemeyer, M. et al. Immunolocalization of the Nuk receptor tyrosine kinase suggests roles in segmental patterning of the brain and axonogenesis. Oncogene 9, 1001–1014 (1994).

    CAS  PubMed  Google Scholar 

  38. Dalva, M.B. et al. EphB receptors interact with NMDA receptors and regulate excitatory synapse formation. Cell 103, 945–956 (2000).

    Article  CAS  Google Scholar 

  39. Jones, T.A., Zou, J.Y. & Cowan, S.W. & Kjeldgaard, M. Improved methods for building protein models in electron density maps and the location of errors in these models. Acta Crystallogr. A 47, 110–119 (1991).

    Article  Google Scholar 

  40. Otwinowski, Z. & Minor, W. Processing of X-ray diffraction data collected in oscillation mode. Methods Enzymol. 276, 307–326 (1997).

    Article  CAS  Google Scholar 

  41. CCP4. The CCP4 suite: programs for X-ray crystallography. Acta Crystallogr. D 50, 760–763 (1994).

  42. Brunger, A.T. et al. Crystallography & NMR system: a new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank M. Greenberg for the gift of phospho-EphB2 antibodies. This work was supported by the US National Institutes of Health (RO1-NS38486 to D.B.N. and RO1-MH66332 to M.H.) and by the Christopher Reeve Paralysis Foundation (to M.J.C. and M.H.). M.H. is a Rita Allen Foundation Scholar.

Author information

Author notes

  1. Martin Lackmann & Christopher Vearing

    Present address: Department of Biochemistry and Molecular Biology, Monash University, Clayton, Melbourne, Victoria, 3168, Australia

  2. Juha-Pekka Himanen and Michael J Chumley: These authors contributed equally to this work.

Authors and Affiliations

  1. Structural Biology Program, Memorial Sloan-Kettering Cancer Center, New York, 10021, New York, USA

    Juha-Pekka Himanen, Chen Li, William A Barton, Phillip D Jeffrey & Dimitar B Nikolov

  2. Center for Developmental Biology and Kent Waldrep Center for Basic Research on Nerve Growth and Regeneration, University of Texas Southwestern Medical Center, Dallas, 75390-9133, Texas, USA

    Michael J Chumley & Mark Henkemeyer

  3. Ludwig Institute for Cancer Research, P.O. Royal Melbourne Hospital, Victoria, 3050, Victoria, Australia

    Martin Lackmann, Christopher Vearing & Detlef Geleick

  4. Department of Molecular, Cellular, and Developmental Biology, University of California at Santa Cruz, Santa Cruz, 95064, California, USA

    David A Feldheim

  5. Queensland Institute of Medical Research, PO Royal Brisbane Hospital, Queensland, 4029, Australia

    Andrew W Boyd

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  1. Juha-Pekka Himanen
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Correspondence to Dimitar B Nikolov.

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Supplementary information

Supplementary Video 1

NG108 cells are unresponsive to pre-clustered ephrin-B1-Fc. (MOV 495 kb)

Supplementary Video 2

Growth cone collapse and neurite retraction in NG108-EphB2 expressing cells following treatment with pre-clustered ephrin-B1-Fc. (MOV 529 kb)

Supplementary Video 3

NG108-EphB2 expressing cells are unresponsive to pre-clustered ephrin-A1-Fc. (MOV 459 kb)

Supplementary Video 4

Growth cone collapse and neurite retraction in NG108-EphB2 expressing cells following treatment with pre-clustered ephrin-A5-Fc. (MOV 491 kb)

Supplementary Video 5

Growth cone collapse in NG108-EphB2 expressing cells following treatment with pre-clustered ephrin-A5-Fc. (MOV 834 kb)

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Himanen, JP., Chumley, M., Lackmann, M. et al. Repelling class discrimination: ephrin-A5 binds to and activates EphB2 receptor signaling. Nat Neurosci 7, 501–509 (2004). https://doi.org/10.1038/nn1237

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