Nucleic Acids Research Advance Access originally published online on December 20, 2006
Nucleic Acids Research 2007 35(3):740-754; doi:10.1093/nar/gkl1053
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Nucleic Acids Research, 2007, Vol. 35, No. 3 740-754
© 2006 The Author(s).
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Nucleic Acid Enzymes |
Specific targeting of cytosine methylation to DNA sequences in vivo
King's College London, Department of Haematological and Molecular Medicine, Guy's, King's and St Thomas' School of Medicine, The Rayne Institute 123 Cold Harbour Lane, London SE5 9NU, UK
*To whom correspondence should be addressed. Tel: +44 207 848 5909; Fax: +44 207 733 3877; Email: kevin.ford{at}kcl.ac.uk
Received July 20, 2006. Revised October 31, 2006. Accepted November 19, 2006.
Development of methods that will allow exogenous imposition of inheritable gene-specific methylation patterns has potential application in both therapeutics and in basic research. An ongoing approach is the use of targeted DNA methyltransferases, which consist of a fusion between gene-targeted zinc-finger proteins and prokaryotic DNA cytosine methyltransferases. These enzymes however have so far demonstrated significant and unacceptable levels of non-targeted methylation. We now report the development of second-generation targeted methyltransferase enzymes comprising enhanced zinc-finger arrays coupled to methyltransferase mutants that are functionally dominated by their zinc-finger component. Both in vitro plasmid methylation studies and a novel bacterial assay reveal a high degree of target-specific methylation by these enzymes. Furthermore, we demonstrate for the first time transient expression of targeted cytosine methyltransferase in mammalian cells resulting in the specific methylation of a chromosomal locus. Importantly, the resultant methylation pattern is inherited through successive cell divisions.
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