Two distinct domains in mouse IG-DMR of the Dlk1–Dio3 locus confer imprinted DNA methylation in transgenic mice

Abstract

Abstract Faithful maintenance of allele-specific methylation at imprinting control regions (ICR) is essential for genomic imprinting. Using transgenic mice (TgM), we investigated sequence requirements for paternal methylation acquisition and maternal hypomethylation maintenance during early embryogenesis. We previously showed that a 2.9-kb mouse H19 ICR acquires post-fertilization paternal allele-specific DNA methylation via a critical 118-bp region; its deletion from the endogenous H19 ICR abolishes maintenance of germ cell-established paternal methylation. In contrast, TgM with the 2.8-kb rat H19 ICR ortholog failed to acquire post-fertilization imprinted methylation. Although another paternally methylated ICR, the IG-DMR of the Dlk1–Dio3 locus, acquired paternal methylation post-fertilization in TgM, the imprinted state was lost due to maternal de novo methylation post-implantation. Here, we extended previously analyzed ICR sequences to reexamine conservation of the activity across different species and genomic loci. Extending the rat H19 ICR upstream to include candidate cis -motifs still failed to establish imprinted methylation. Conversely, TgM with an extended mouse IG-DMR sequence protected maternal IG-DMR from genome-wide de novo DNA methylation after implantation, maintaining its imprinted DNA methylation state. This finding provides the first in vivo evidence for a bipartite IG-DMR model: 5′ domain for paternal methylation acquisition and 3′ domain for maternal hypomethylation maintenance.