RT Journal Article SR Electronic T1 A bipartite element with allele-specific functions safeguards DNA methylation imprints at the Dlk1-Dio3 locus JF bioRxiv FD Cold Spring Harbor Laboratory SP 2020.05.22.103796 DO 10.1101/2020.05.22.103796 A1 BE Aronson A1 L Scourzic A1 V Shah A1 E Swanzey A1 A Kloetgen A1 A Polyzos A1 A Sinha A1 A Azziz A1 I Caspi A1 J Li A1 B Pelham-Webb A1 H Wichterle A1 A Tsirigos A1 M Stadtfeld A1 E Apostolou YR 2020 UL http://biorxiv.org/content/early/2020/05/24/2020.05.22.103796.abstract AB Dysregulation of imprinted gene loci also referred to as loss of imprinting (LOI) can result in severe developmental defects and other diseases, but the molecular mechanisms that ensure imprint stability remain incompletely understood. Here, we dissect the functional components of the imprinting control region of the essential Dlk1-Dio3 locus (called IG-DMR) and the mechanism by which they ensure imprinting maintenance. Using pluripotent stem cells carrying an allele-specific reporter system, we demonstrate that the IG-DMR consists of two antagonistic regulatory elements: a paternally methylated CpG-island that prevents the activity of Tet dioxygenases and a maternally unmethylated regulatory element, which serves as a non-canonical enhancer and maintains expression of the maternal Gtl2 lncRNA by precluding de novo DNA methyltransferase function. Targeted genetic or epigenetic editing of these elements leads to LOI with either bi-paternal or bi-maternal expression patterns and respective allelic changes in DNA methylation and 3D chromatin topology of the entire Dlk1-Dio3 locus. Although the targeted repression of either IG-DMR or Gtl2 promoter is sufficient to cause LOI, the stability of LOI phenotype depends on the IG-DMR status, suggesting a functional hierarchy. These findings establish the IG-DMR as a novel type of bipartite control element and provide mechanistic insights into the control of Dlk1-Dio3 imprinting by allele-specific restriction of the DNA (de)methylation machinery.HIGHLIGHTSThe IG-DMR is a bipartite element with distinct allele-specific functionsA non-canonical enhancer within the IG-DMR prevents DNA methyltransferase activityTargeted epigenome editing allows induction of specific imprinting phenotypesCRISPRi reveals a functional hierarchy between DMRs that dictates imprint stabilityCompeting Interest StatementThe authors have declared no competing interest.