BMP inhibition-driven regulation of six-3 underlies induction of newt lens regeneration

Abstract

Lens regeneration in adult newts is a classic example of how cells can faithfully regenerate a complete organ through the process of transdifferentiation1,2,3,4,5,6. After lens removal, the pigment epithelial cells of the dorsal, but not the ventral, iris dedifferentiate and then differentiate to form a new lens. Understanding how this process is regulated might provide clues about why lens regeneration does not occur in higher vertebrates. The genes six-3 and pax-6 are known to induce ectopic lenses during embryogenesis7,8. Here we tested these genes, as well as members of the bone morphogenetic protein (BMP) pathway that regulate establishment of the dorsal–ventral axis in embryos9, for their ability to induce lens regeneration. We show that the lens can be regenerated from the ventral iris when the BMP pathway is inhibited and when the iris is transfected with six-3 and treated with retinoic acid. In intact irises, six-3 is expressed at higher levels in the ventral than in the dorsal iris. During regeneration, however, only expression in the dorsal iris is significantly increased. Such an increase is seen in ventral irises only when they are induced to transdifferentiate by six-3 and retinoic acid or by BMP inhibitors. These data suggest that lens regeneration can be achieved in noncompetent adult tissues and that this regeneration occurs through a gene regulatory mechanism that is more complex than the dorsal expression of lens regeneration-specific genes.

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Figure 1: Lens induction from ventral PECs.
Figure 2: Expression during lens regeneration and induction.
Figure 3: Expression of Six-3 and BMPR-IA during regeneration.
Figure 4: Expression in axolotl.

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Acknowledgements

We thank T. Hayashi for sharing his expertise with the culturing methods for PECs; G. Oliver and D. Englecamp for the six-3 and pax-6 expression vectors, respectively; members of our laboratories, M. Metheney, A. Thitoff, S. Gainer, M. Tsavaris, M. Tubb and M. Sander for help with histology; V. Mitashov and E. Makarev for providing information on P.Waltl six-3 sequences; and E. Tanaka for information on axolotl BMPR-IA sequences. This work was supported by a grant from the NIH (to P.A.T.).

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Correspondence to Panagiotis A. Tsonis.

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Competing interests

Sequences for six-3 and BMPR-IA have been deposited in GenBank under accession numbers AY799802 and AY795966, respectively. Reprints and permissions information is available at npg.nature.com/reprintsandpermissions. The authors declare no competing financial interests.

Supplementary information

Supplementary Table 1

Describes the cases of induction of lens regeneration in PEC aggregates or explants in response to six-3 transfections and BMP treatments. (DOC 34 kb)

Supplementary Figure 1

Shows the efficiency of transfection and the expression of exogenous genes. (PDF 1448 kb)

Supplementary Figure Legend

Text to accompany the above Supplementary Figure. (DOC 24 kb)

Supplementary Methods

Several methods on the isolation, culture, treatment and transfection of PECs. (DOC 30 kb)

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Grogg, M., Call, M., Okamoto, M. et al. BMP inhibition-driven regulation of six-3 underlies induction of newt lens regeneration. Nature 438, 858–862 (2005). https://doi.org/10.1038/nature04175

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