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Functional gene screening in embryonic stem cells implicates Wnt antagonism in neural differentiation

Nature Biotechnology volume 20, pages 1240–1245 (2002)Cite this article

Abstract

The multilineage differentiation capacity of mouse embryonic stem (ES) cells offers a potential testing platform for gene products that mediate mammalian lineage determination and cellular specialization. Identification of such differentiation regulators is crucial to harnessing ES cells for pharmaceutical discovery and cell therapy. Here we describe the use of episomal expression technology for functional evaluation of cDNA clones during ES-cell differentiation in vitro. Several candidate cDNAs identified by subtractive cloning and expression profiling were introduced into ES cells in episomal expression constructs. Subsequent differentiation revealed that the Wnt antagonist Sfrp2 stimulates production of neural progenitors. The significance of this observation was substantiated by forced expression of Wnt-1 and treatment with lithium chloride, both of which inhibit neural differentiation. These findings reveal the importance of Wnt signaling in regulating ES-cell lineage diversification. More generally, this study establishes a path for rapid and direct validation of candidate genes in ES cells.

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Figure 1: Experimental overview.
Figure 2: Expression of selected cDNAs in the embryonic CNS.
Figure 3: Effect of Sfrp2 expression in E14/T cells during ES cell differentiation.
Figure 4: Effect of forced expression of Sfrp2 on differentiation of Sox1-GFP ES cells.
Figure 5: Wnt activity suppresses neural differentiation.

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Acknowledgements

We thank Marios Stavridis for 46C ES cells, and Meng Li, Jenny Nichols, Tom Burdon, and Joe Mee for advice and discussion. This research was supported by the Medical Research Council and the Biotechnology and Biological Sciences Research Council of the UK. J.A. was supported by an INSERM fellowship and a Marie Curie EU Fellowship.

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  1. Institute for Stem Cell Research, University of Edinburgh, The King's Buildings, West Mains Road, Edinburgh, EH3 9JQ, UK

    Jerôme Aubert, Hannah Dunstan, Ian Chambers & Austin Smith

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  1. Jerôme Aubert
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Correspondence to Austin Smith.

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A.S. is a scientific advisor to Stem Cell Sciences Ltd. and holds non-voting equity in the company. Stem Cell Sciences funds research in the laboratory and has patents pending on technology used in the work described in this article.

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Aubert, J., Dunstan, H., Chambers, I. et al. Functional gene screening in embryonic stem cells implicates Wnt antagonism in neural differentiation. Nat Biotechnol 20, 1240–1245 (2002). https://doi.org/10.1038/nbt763

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