GERM-LINE PASSAGE IS REQUIRED FOR ESTABLISHMENT OF METHYLATION AND EXPRESSION PATTERNS OF IMPRINTED BUT NOT OF NONIMPRINTED GENES

Embryonic stem (ES) cells homozygous for a disruption of the DNA (cyto sine-5)-methyltransferase gene (Dnmt) proliferate normally with their DNA highly demethylated but die upon differentiation. Expression of th e wild-type Dnmt cDNA in mutant male ES cells caused an increase in me thylation 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 Ig f2r. These cells differentiated normally in vitro and contributed subs tantially to adult chimeras. While the Xist gene was not expressed in the remethylated male ES cells, no restoration of the normal expressio n profile was seen for H19, Igf2r, or Igf2. This indicates that ES cel ls can faithfully reestablish normal methylation and expression patter ns of nonimprinted genes but lack the ability to restore those of impr inted genes. Full restoration of monoallelic methylation and expressio n was imposed on H19, Igf2, and Igf2r upon germ-line transmission. The se results are consistent with the presence of distinct de novo DNA me thyltransferase activities during oogenesis and spermatogenesis, which specifically recognize imprinted genes but are absent in the postimpl antation embryo and in ES cells.