DNA Demethylation Reactivates a Subset of Imprinted Genes in Uniparental Mouse Embryonic Fibroblasts

Abstract

Although most imprinted genes show allelic differences in DNA methylation, it is not clear whether methylation regulates the expression of some or all imprinted genes in somatic cells. To examine the mechanisms of silencing of imprinted alleles, we generated novel uniparental mouse embryonic fibroblasts exclusively containing either the paternal or the maternal genome. These fibroblasts retain parent-of-origin allele-specific expression of 12 imprinted genes examined for more than 30 cell generations. We show that p57(Kip2) (cyclin-dependent kinase inhibitor protein 2) and Igf2 (insulin-like growth factor 2) are induced by inhibiting histone deacetylases; however, their activated state is reversed quickly by withdrawal of trichostatin A. In contrast, DNA demethylation results in the heritable expression of a subset of imprinted genes including H19 (H19 fetal liver mRNA), p57(Kip2), Peg3/Pw1 (paternally expressed gene 3), and Zac1 (zinc finger-binding protein regulating apoptosis and cell cycle arrest). Other imprinted genes such as Grb10 (growth factor receptor-bound protein 10), Peg1/Mest (paternally expressed gene 1/mesoderm-specific transcript), Sgce (epsilon-sarcoglycan), Snrpn (small nuclear ribonucleoprotein polypeptide N), and U2af1 (U2 small nuclear ribonucleoprotein auxiliary factor), remain inactive, despite their exposure to inhibitors of histone deacetylases and DNA methylation. These results demonstrate that changes in DNA methylation but not histone acetylation create a heritable epigenetic state at some imprinted loci in somatic cells.