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DNA demethylation is necessary for the epigenetic reprogramming of somatic cell nuclei

Nature Cell Biology volume 6, pages 984–990 (2004)Cite this article

Abstract

Nuclear transplantation experiments in amphibia and mammals have shown that oocyte and egg cytoplasm can extensively reprogram somatic cell nuclei with new patterns of gene expression and new pathways of cell differentiation1,2,3; however, very little is known about the molecular mechanism of nuclear reprogramming. Here we have used nuclear and DNA transfer from mammalian somatic cells to analyse the mechanism of activation of the stem cell marker gene oct4 by Xenopus oocytes. We find that the removal of nuclear protein accelerates the rate of reprogramming, but even more important is the demethylation of somatic cell DNA. DNA demethylation seems to precede gene reprogramming, and is absolutely necessary for oct4 transcription. Reprogramming by oocytes occurs in the absence of DNA replication and RNA/protein synthesis. It is also selective, operating only on the promoter, but not enhancers, of oct4; both a putative Sp1/Sp3 and a GGGAGGG binding site are required for demethylation and transcription. We conclude that the demethylation of promoter DNA may be a necessary step in the epigenetic reprogramming of somatic cell nuclei.

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Figure 1: Deproteinization of nuclei and unmethylated DNA accelerate the time when oct4 transcription starts.
Figure 2: The mouse oct4 promoter is demethylated by Xenopus oocytes.
Figure 3: The regulatory region of mouse oct4 in plasmid DNA can be used to analyse the demethylating activity of Xenopus oocytes.
Figure 4: Mutational analysis of oct4 promoter demethylation.

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Acknowledgements

We thank T.J. Mohun, G. Spohr and A. Smith for gifts of DNA constructs; N. Garrett for help, especially with the construction of DNA mutations; C. Lee for a supply of mouse thymocytes; and P. Hurd, A. Bannister, P. Hajkova and J. Byrne for discussion. Our work is supported by the Biotechnology and Biological Sciences Research Council, the Swedish Foundation for International Cooperation in Research and Higher Education, the Wellcome Trust and the European Commission.

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  1. Wellcome Trust/Cancer Research UK Gurdon Institute and Department of Zoology, University of Cambridge, Cambridge, CB2 1QR, UK

    Stina Simonsson & John Gurdon

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  1. Stina Simonsson
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  2. John Gurdon
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Correspondence to John Gurdon.

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Simonsson, S., Gurdon, J. DNA demethylation is necessary for the epigenetic reprogramming of somatic cell nuclei. Nat Cell Biol 6, 984–990 (2004). https://doi.org/10.1038/ncb1176

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