Genomic imprinting is a major monoallelic gene expression regulatory mechanism in mammals, and depends on gamete-specific DNA methylation of specialized cis-regulatory elements called imprinting control regions (ICRs). Allele-specific DNA methylation of the ICRs is faithfully maintained at the imprinted loci throughout development, even in early embryos where genomes undergo extensive epigenetic reprogramming, including DNA demethylation, to acquire totipotency. We previously found that an ectopically introduced H19 ICR fragment in transgenic mice acquired paternal allele-specific methylation in the somatic cells of offspring, while it was not methylated in sperm, suggesting its gametic and postfertilization modifications are separable events. We hypothesized that this latter activity might contribute to maintenance of the methylation imprint in early embryos. Here we demonstrate that methylation of the paternally inherited transgenic H19 ICR commences soon after fertilization in a maternal Dnmt3a- and Dnmt3L-dependent manner. When its germline methylation was partially obstructed by insertion of insulator sequences, the endogenous, paternal H19 ICR also exhibited postfertilization methylation. Finally, we refined the responsible sequences for this activity in transgenic mice, and found that deletion of the 5' segment of the endogenous paternal H19 ICR decreased its methylation after fertilization, attenuated Igf2 gene expression. These results demonstrate that this segment of the H19 ICR is essential for its de novo post-fertilization DNA methylation, and that this activity contributes to the maintenance of imprinted methylation at the endogenous H19 ICR during early embryogenesis
- Received May 2, 2015.
- Accepted September 15, 2015.