The following is the abstract from a paper recently accepted for publication in Epigenetics & Chromatin.
Asymmetric DNA methylation of CpG dyads is a feature of secondary DMRs associated with the Dlk1/Gtl2 imprinting cluster in mouse.
Megan Guntrum (’16), Ekaterina Vlasova (’15) and Tamara L. Davis
Background: Differential DNA methylation plays a critical role in the regulation of imprinted genes. The differentially methylated state of the imprinting control region is inherited via the gametes at fertilization, and is stably maintained in somatic cells throughout development, influencing the expression of genes across the imprinting cluster. In contrast, DNA methylation patterns are more labile at secondary differentially methylated regions which are established at imprinted loci during post-implantation development. To investigate the nature of these more variably methylated secondary differentially methylated regions, we adopted a hairpin-linker bisulfite mutagenesis approach to examine CpG dyad methylation at differentially methylated regions associated with the murine Dlk1/Gtl2 imprinting cluster on both complementary strands. Results: We observed homomethylation at greater than 90% of the methylated CpG dyads at the IG-DMR, which serves as the imprinting control element. In contrast, homomethylation was only observed at 67-78% of the methylated CpG dyads at the secondary differentially methylated regions; the remaining 22-33% of methylated CpG dyads exhibited hemimethylation. Conclusion: We propose that this high degree of hemimethylation could explain the variability in DNA methylation patterns at secondary differentially methylated regions associated with imprinted loci. We further suggest that the presence of 5-hydroxymethylation at secondary differentially methylated regions may result in hemimethylation and methylation variability as a result of passive and/or active demethylation mechanisms.
The following is the abstract for a poster presented at the Gordon Research Conference on Epigenetics, August 2-7, 2015.
Analysis of hemimethylation & 5hmC content at primary and secondary DMRs associated with imprinted loci
Ekaterina Vlasova (’15), Megan Guntrum (’16), Jessica Arbon (’14) and Tamara L. Davis
Differential distribution of DNA methylation on the alleles of imprinted genes functions both to distinguish the alleles based on their parental origin and regulate them to achieve monoallelic expression. DNA methylation at primary differentially methylated regions (DMRs), or imprinting control regions (ICRs) is inherited from the gametes, is consistently maintained on one parental allele throughout development, and functions to modulate expression. In contrast, parent of origin-specific DNA methylation at secondary DMRs is acquired during post-fertilization development; while DNA methylation of these regions is believe to play a role in maintaining imprinted expression, they lack the stability of DNA methylation that is inherited via the gamete. Our analysis of the variable DNA methylation pattern at the mouse Dlk1 gene identified high levels of hemimethylation at individual CpG dyads. We propose that increased levels of hemimethylation at secondary DMRs contribute to the reduced DNA methylation fidelity within these regions. We further propose that increases in 5-hydroxymethylcytosine (5hmC) levels may be responsible for increases in hemimethylation and decreases in DNA methylation fidelity. To test these hypotheses, we are examining CpG dyad methylation patterns and 5hmC content at both primary and secondary DMRs associated with imprinted genes. This poster will describe our recent results and current investigations.