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Rac-dependent trans-endocytosis of ephrinBs regulates Eph–ephrin contact repulsion

Nature Cell Biology volume 5, pages 879–888 (2003)Cite this article

Abstract

Eph receptor–ephrin signals are important for controlling repulsive and attractive cell movements during tissue patterning in embryonic development. However, the dynamic cellular responses to these signals at cell–cell contact sites are poorly understood. To examine these events we have used cell microinjection to express EphB4 and ephrinB2 in adjacent Swiss 3T3 fibroblasts and have studied the interaction of the injected cells using time-lapse microscopy. We show that Eph receptors are locally activated wherever neighbouring cells make contact. This triggers dynamic, Rac-regulated membrane ruffles at the Eph–ephrin contact sites. Subsequently, the receptor and ligand cells retract from one another, concomitantly with the endocytosis of the activated Eph receptors and their bound, full-length ephrinB ligands. Both the internalization of the receptor–ligand complexes and the subsequent cell retraction events are dependent on actin polymerization, which in turn is dependent on Rac signalling within the receptor-expressing cells. Similar events occur in primary human endothelial cells. Our findings suggest a novel mechanism for cell repulsion, in which the contact between Eph-expressing and ephrin-expressing cells is destabilized by the localized phagocytosis of the ligand-expressing cell plasma membrane by the receptor-expressing cell.

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Figure 1: Activation of EphB4 at ephrinB2 cell contact sites.
Figure 2: Local activation of EphB4 and ephrinB2 induces membrane ruffling and cell retraction.
Figure 3: Endocytosis of p-EphB4 from Eph–ephrin activation sites.
Figure 4: Full-length ephrinB is transferred to EphB4-expressing cells.
Figure 5: Lamellipodia and filopodia extend from activated receptor cells.
Figure 6: Retraction and endocytosis require actin polymerization.
Figure 7: Rac is required for ephrinB2-induced retraction of endothelial cells.
Figure 8: Model of contact repulsion regulated by ephrinB trans-endocytosis.

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Acknowledgements

We thank R. Klein for sharing unpublished results. We also thank R. Adams, D. Wilkinson, M. Henkemeyer, C. Cowan, D. Holowka, L. Machesky, M. Furthmayr and J. M. Edwardson for cDNAs and expression constructs; J. Pendjiky, D. Ciantar and M. Shipman for help with image acquisition and processing; P. Martin and M. Raff for valuable comments on the manuscript, and members of the Nobes laboratory for advice and helpful discussions. This work was inspired by the late Nigel Holder and was supported by the Medical Research Council. C.D.N. is an MRC Senior Research Fellow.

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  1. Sarah Dickinson & Catherine D. Nobes

    Present address: Department of Physiology and Biochemistry, School of Medical Sciences, University of Bristol, Bristol, BS8 1TD, UK

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  1. Department of Anatomy and Developmental Biology, Centre for Cell and Molecular Dynamics, University College London, London, WC1E 6BT, UK

    Daniel J. Marston, Sarah Dickinson & Catherine D. Nobes

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Correspondence to Catherine D. Nobes.

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Marston, D., Dickinson, S. & Nobes, C. Rac-dependent trans-endocytosis of ephrinBs regulates Eph–ephrin contact repulsion. Nat Cell Biol 5, 879–888 (2003). https://doi.org/10.1038/ncb1044

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