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The transcription factor Engrailed-2 guides retinal axons

Nature volume 438, pages 94–98 (2005)Cite this article

Abstract

Engrailed-2 (En-2), a homeodomain transcription factor, is expressed in a caudal-to-rostral gradient in the developing midbrain, where it has an instructive role in patterning the optic tectum—the target of topographic retinal input1,2. In addition to its well-known role in regulating gene expression through its DNA-binding domain, En-2 may also have a role in cell–cell communication, as suggested by the presence of other domains involved in nuclear export, secretion and internalization3. Consistent with this possibility, here we report that an external gradient of En-2 protein strongly repels growth cones of Xenopus axons originating from the temporal retina and, conversely, attracts nasal axons. Fluorescently tagged En-2 accumulates inside growth cones within minutes of exposure, and a mutant form of the protein that cannot enter cells fails to elicit axon turning. Once internalized, En-2 stimulates the rapid phosphorylation of proteins involved in translation initiation and triggers the local synthesis of new proteins. Furthermore, the turning responses of both nasal and temporal growth cones in the presence of En-2 are blocked by inhibitors of protein synthesis. The differential guidance of nasal and temporal axons reported here suggests that En-2 may participate directly in topographic map formation in the vertebrate visual system.

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Figure 1: En-2 gradient repels temporal and attracts nasal retinal axons.
Figure 2: En-2 is internalized in the growth cone and guidance depends on internalization.
Figure 3: Retinal ganglion cell growth cone guidance by En-2 is dependent on protein synthesis.
Figure 4: En-2 stimulates the phosphorylation of translational regulatory proteins.

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Acknowledgements

We thank all of our colleagues for discussions, and A. Dwivedy and E. Ipendey for technical assistance. This work was supported by the Wellcome Trust, UK (to C.H. and W.H.), the Human Frontier Sciences Program and the European Commission (to A.P.). Author Contributions I.B. did the turning assays and internalization experiments in Figs 1, 2, 3a and 3c, and in the Supplementary Figs. C.W. did the quantitative immunofluorescence work in Fig. 4, and trained I.B. in turning assays. M.P. did the 3H-leucine experiments in Fig. 3b. M.V. provided all the constructs and proteins. A.P. proposed the hypothesis that En-2 guides retinal axons. A.P. and A.T. initiated the collaboration and discussed experiments. W.H. and C.H. wrote the manuscript and discussed the experiments.

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Authors and Affiliations

  1. CNRS UMR 8542, Ecole Normale Supérieure, 46 rue d'Ulm, 75230, Paris, Cedex 05, France

    Isabelle Brunet, Alain Trembleau, Michel Volovitch & Alain Prochiantz

  2. Department of Anatomy, University of Cambridge, Downing Street, CB2 3DY, Cambridge, UK

    Isabelle Brunet, Christine Weinl, Michael Piper, William Harris & Christine Holt

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  1. Isabelle Brunet
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Correspondence to Alain Prochiantz or Christine Holt.

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Supplementary information

Supplementary Figures

Supplementary Figures 1–4 show that En-2 induces opposite turning behaviour in nasal and temporal axons in ‘blind’ experiments and does not alter their extension rate. Photomicrographs show growth cones severed from their cell bodies exhibit differential turning in an En-2 gradient. Otx2 only elicits weakly attractive turning responses. (DOC 526 kb)

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Brunet, I., Weinl, C., Piper, M. et al. The transcription factor Engrailed-2 guides retinal axons. Nature 438, 94–98 (2005). https://doi.org/10.1038/nature04110

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