Research article

Direct non-cell autonomous Pax6 activity regulates eye development in the zebrafish

Brigitte Lesaffre¹ , Alain Joliot² , Alain Prochiantz¹ and Michel Volovitch¹

¹  Development and Neuropharmacology, CNRS UMR 8542 and Ecole Normale Supérieure, rue d'Ulm, 75230 Paris Cedex 05, France

²  Homeoprotein cell biology, CNRS UMR 8542 and Ecole Normale Supérieure, rue d'Ulm, 75230 Paris Cedex 05, France

author email corresponding author email

Neural Development 2007, 2:2doi:10.1186/1749-8104-2-2

Published:	17 January 2007

Abstract

Background

Modifications in Pax6 homeogene expression produce strong eye phenotypes. This suggested to us that eye development might be an appropriate model to verify if homeoprotein intercellular passage has important functions in early development. Similar to other homeoproteins, Pax6 has two domains that enable secretion and internalization by live cells and, thus, intercellular passage. In principle, a straightforward way to test the hypothesis would be to mutate one of the two sequences to produce a 'cell autonomous only' Pax6. However, this was not possible because these sequences are in the homeodomain and their modification would affect Pax6 transcriptional properties. We have thus developed an approach aimed at blocking Pax6 only in the extracellular milieu of developing zebrafish embryos.

Results

A first strategy was to inject a one-cell embryo with a mRNA encoding a secreted single-chain anti-Pax6 antibody. A second, complementary, strategy was to inject a Pax6 antibody in the blastula extracellular milieu. In both cases, 'dissymmetric eyes', 'one eye only' and 'no eye' phenotypes were produced. In most cases, lens phenotypes paralleled retina malformations. Although eye phenotypes were analyzed 30 hours post-fertilization, there was a strong correlation between early eye field asymmetry, early asymmetry in Pax6 expression and later-occurring eye malformations. Several controls were introduced, demonstrating that the effect is specific to Pax6 and cannot be explained by intracellular antibody activities.

Conclusion

This study supports the hypothesis that the Pax6 transcription factor is also a signaling molecule with direct non-cell autonomous activity.