Widespread recovery of methylation at gametic imprints in hypomethylated cells following rescue with DNMT3A2.

Imprinted loci are paradigms of epigenetic regulation and are associated with a number of genetic disorders in human. A key characteristic of the imprints is the presence of a gametic differentially methylated region (gDMR) which must be passaged through the germline to be reprogrammed. We examined the most complete current set of imprinted gDMRs in mouse embryonic stem cells (ESCs) lacking either DNMT1 (1KO) or a combination of DNMT3A and DNMT3B (3abKO) and found that loss of methylation (5mC) was approximately equivalent. 1KO cells rescued with a cDNA expressing DNMT1 could not restore methylation at the imprinted gDMRs, as has previously been shown. Surprisingly however, nearly all gDMRs were remethylated in 3abKO cells rescued with a DNMT3A2 expression construct. These results were confirmed using three different approaches. Transcriptional activity at the H19/Igf2 locus also tracked with the methylation pattern, confirming functional reprogramming. These results suggested 1) a vital role for DNMT3A/B in methylation maintenance at imprints 2) that loss of DNMT1 and DNMT3A/B had equivalent effects 3) that rescue with DNMT3A2 can restore imprints in this model system. This system offers a potentially useful model in which to further explore aspects of imprint reprogramming