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Grb4 and GIT1 transduce ephrinB reverse signals modulating spine morphogenesis and synapse formation

Nature Neuroscience volume 10, pages 301–310 (2007)Cite this article

Abstract

Dendritic spines are small protrusions emerging from dendrites that receive excitatory input. The process of spine morphogenesis occurs both in the developing brain and during synaptic plasticity. Molecules regulating the cytoskeleton are involved in spine formation and maintenance. Here we show that reverse signaling by the transmembrane ligands for Eph receptors, ephrinBs, is required for correct spine morphogenesis. The molecular mechanism underlying this function of ephrinBs involves the SH2 and SH3 domain–containing adaptor protein Grb4 and the G protein–coupled receptor kinase–interacting protein (GIT) 1. Grb4 binds by its SH2 domain to Tyr392 in the synaptic localization domain of GIT1. Phosphorylation of Tyr392 and the recruitment of GIT1 to synapses are regulated by ephrinB activation. Disruption of this pathway in cultured rat hippocampal neurons impairs spine morphogenesis and synapse formation. We thus show an important role for ephrinB reverse signaling in spine formation and have mapped the downstream pathway involved in this process.

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Figure 1: EphrinB signaling promotes spine morphogenesis.
Figure 2: Interference with ephrinB reverse signaling results in impaired spine formation.
Figure 3: Grb4 localizes to synapses.
Figure 4: GIT1 interacts with Grb4.
Figure 5: Grb4-GIT1 binding is regulated by ephrinB reverse signaling.
Figure 6: GIT1 is recruited to ephrinB patches at the synapse.
Figure 7: Disruption of ephrinB signaling through Grb4 and GIT1 affects spine morphogenesis.
Figure 8: Disruption of ephrinB signaling through Grb4 and GIT1 affects synapse formation.

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Acknowledgements

We would like to thank Cellzome AG, A. Senturk, S. Jordan, M. Zimmer, and E. Martinez for technical help; R.T. Premont (Duke University Medical Center) for GIT1- and GIT2-Flag constructs and antibodies to GIT1, GIT2 and β-PIX; K.S. Erdmann (Ruhr-University) for HeLa-ephrinB1 cells; R. Klein for helpful discussions and A. Yamasaki, I. Kadow and R. Klein for critically reading the manuscript. This work was supported by grants from the Deutsche Forschungsgemeinschaft (AC180/2-1 to A.A.-P.).

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Author notes

  1. Inmaculada Segura, Clara L Essmann and Stefan Weinges: These authors contributed equally to this work.

Authors and Affiliations

  1. Max-Planck Institute of Neurobiology, Am Klopferspitz 18, Martinsried, D-82152, Germany

    Inmaculada Segura, Clara L Essmann, Stefan Weinges & Amparo Acker-Palmer

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  1. Inmaculada Segura
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Contributions

I.S. and S.W. performed the molecular biology and biochemical experiments. C.L.E. conducted the spine assays and dominant negative experiments with the help of I.S. and S.W. A.A.-P. designed and supervised the experiments and wrote the manuscript.

Corresponding author

Correspondence to Amparo Acker-Palmer.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Spine maturation is independently driven by Eph receptor forward and ephrinB reverse signaling. (PDF 715 kb)

Supplementary Fig. 2

GIT proteins interact with Grb4 through the SLD. (PDF 1224 kb)

Supplementary Fig. 3

Grb4 interacts with GIT1 and ephrinB ligands though SH2 and SH3 domains. (PDF 2633 kb)

Supplementary Methods (PDF 117 kb)

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Segura, I., Essmann, C., Weinges, S. et al. Grb4 and GIT1 transduce ephrinB reverse signals modulating spine morphogenesis and synapse formation. Nat Neurosci 10, 301–310 (2007). https://doi.org/10.1038/nn1858

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