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An evolutionary recent neuroepithelial cell adhesion function of huntingtin implicates ADAM10-Ncadherin

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Abstract

The Huntington's disease gene product, huntingtin, is indispensable for neural tube formation, but its role is obscure. We studied neurulation in htt-null embryonic stem cells and htt-morpholino zebrafish embryos and found a previously unknown, evolutionarily recent function for this ancient protein. We found that htt was essential for homotypic interactions between neuroepithelial cells; it permitted neurulation and rosette formation by regulating metalloprotease ADAM10 activity and Ncadherin cleavage. This function was embedded in the N terminus of htt and was phenocopied by treatment of htt knockdown zebrafish with an ADAM10 inhibitor. Notably, in htt-null cells, reversion of the rosetteless phenotype occurred only with expression of evolutionarily recent htt heterologues from deuterostome organisms. Conversely, all of the heterologues that we tested, including htt from Drosophila melanogaster and Dictyostelium discoideum, exhibited anti-apoptotic activity. Thus, anti-apoptosis may have been one of htt's ancestral function(s), but, in deuterostomes, htt evolved to acquire a unique regulatory activity for controlling neural adhesion via ADAM10-Ncadherin, with implications for brain evolution and development.

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Figure 1: Neural differentiation of htt-null mouse ES cells.
Figure 2: Neural tube impairment in htt loss-of-function zebrafish embryos.
Figure 3: Lack of htt causes increased ADAM10 activity and Ncadherin cleavage.
Figure 4: Ncadherin cleavage and ADAM10 activity in the zebrafish and mouse brain.
Figure 5: Effects of htt loss-of-function during neurulation.
Figure 6: The block of ADAM10 activity partially rescued httMO phenotypes at 24 hpf.
Figure 7: htt N terminus neural cell-adhesion function during evolution.

Change history

  • 10 August 2012

    In the version of this article initially published, the htt ATG morpholino sequence given in Online Methods (ATTTTAACAGAAGCTGTGATG) was incorrect. The correct sequence is 5′-GCCATTTTAACAGAAGCTGTGATGA-3′. The error has been corrected in the HTML and PDF versions of the article.

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Acknowledgements

We thank M. MacDonald (Massachusetts General Hospital) for the Hdh+/+ and Hdhex4–5 cells, P. Saftig for the ADAM10 antibody (University of Kiel), E. Ruthazer for the zebrafish Ncadherin antibody (McGill University), F. Gardoni and E. Marcello for helpful technical advice on the use of ADAM10 and SAP97 antibodies, A. Badaloni for help with subcloning, and G. Simonutti for assistance with initial imaging analysis. This work has progressed at a very slow pace because it was mostly unfunded. In the beginning, this work was partially supported by the Huntington's Disease Society of America Coalition for the Cure (2007–2009) and by Italian Telethon Foundation (GGP06250, 2007–2009). Some additional support was provided by the Ministero dell'Istruzione, dell'Università e della Ricerca Scientifica, Programmi di Ricerca Scientifica di Rilevante Interesse Nazionale (2006052993, 2006). Since 2009, progress of this work has relied entirely on occasional donations. We wish to thank one donor that in 2009 collected a considerable amount of small donations for us to continue this work and another donor that has been following our progress for 10 years. C.R. was supported by the Fundação para a Ciência e Tecnologia (grant number SFRH/BD/9627/2002) through the GABBA Programme (University of Porto). E.C., J.G. and S.Z. were members of the Huntington's Disease Society of America Coalition for the Cure and J.G. received support from NS16367.

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Authors

Contributions

E.C., V.L.S. and C.Z. developed the study, conceived the experimental plans and analyzed the data. V.L.S. and C.Z. performed most of the biological, biochemical and molecular experiments. B.V. performed some of the biological and biochemical experiments. M.T. and C.R. participated in the initial elaboration of the project and conducted some experiments (some of the initial constructs preparation and monolayer assays, respectively). V.L.S. prepared additional constructs. M.A.M., J.A.W. and J.G. provided the Dictyostelium and Drosophila cDNA. G.G., C.Z. and A.P. performed experiments in zebrafish under the supervision of F.C. (immunocytochemistry and in situ were performed by G.G. and A.P., biochemical assays by C.Z.). M.V. and L.C. provided suggestions for some biological experiments. S.Z. provided the conditional knockout mice and some constructs. B.D. and B.S. provided GI254023X. V.L.S., C.Z. and E.C. interpreted the data and wrote the manuscript. All of the authors read and edited the manuscript. E.C. supervised the entire work, directed the strategies, provided financial support and gave final approval of the version to be published.

Corresponding author

Correspondence to Elena Cattaneo.

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Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–10 (PDF 19081 kb)

Supplementary Video 1

Time lapse Imaging experiment in Hdh+/+ cells during neural differentiation. (MOV 14150 kb)

Supplementary Video 2

Time lapse Imaging experiment in Hdhex4/5 cells during neural differentiation. (MOV 14455 kb)

Supplementary Video 3

Time lapse Imaging experiment of co-culture system during neural differentiation. (MOV 21124 kb)

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Lo Sardo, V., Zuccato, C., Gaudenzi, G. et al. An evolutionary recent neuroepithelial cell adhesion function of huntingtin implicates ADAM10-Ncadherin. Nat Neurosci 15, 713–721 (2012). https://doi.org/10.1038/nn.3080

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