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Research Papers
Whitehead Institute for Biomedical Research, Cambridge, Massachusetts 02142, USA.
Abstract
Embryonic stem (ES) cells homozygous for a disruption of the DNA (cytosine-5)-methyltransferase gene (Dnmt) proliferate normally with their DNA highly demethylated but die upon differentiation. Expression of the wild-type Dnmt cDNA in mutant male ES cells caused an increase in methylation of bulk DNA and of the Xist and Igf2 genes to normal levels, but did not restore the methylation of the imprinted genes H19 and Igf2r. These cells differentiated normally in vitro and contributed substantially to adult chimeras. While the Xist gene was not expressed in the remethylated male ES cells, no restoration of the normal expression profile was seen for H19, Igf2r, or Igf2. This indicates that ES cells can faithfully reestablish normal methylation and expression patterns of nonimprinted genes but lack the ability to restore those of imprinted genes. Full restoration of monoallelic methylation and expression was imposed on H19, Igf2, and Igf2r upon germ-line transmission. These results are consistent with the presence of distinct de novo DNA methyltransferase activities during oogenesis and spermatogenesis, which specifically recognize imprinted genes but are absent in the postimplantation embryo and in ES cells.
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