Imprinted gene expression at the Dlk1-Dio3 cluster is controlled by both maternal and paternal IG-DMRs in a tissue-specific fashion.

Imprinting at the Dlk1-Dio3 cluster is controlled by the IG-DMR, an imprinting control region differentially methylated between maternal and paternal chromosomes. The maternal IG-DMR is essential for imprinting control, functioning as a cis enhancer element. Meanwhile, DNA methylation at the paternal IG-DMR is thought to prevent enhancer activity. To explore whether suppression of enhancer activity at the methylated IG-DMR requires the transcriptional repressor TRIM28, we analyzed Trim28chatwo embryos and performed epistatic experiments with IG-DMR deletion mutants. We found that while TRIM28 regulates the enhancer properties of the paternal IG-DMR, it also controls imprinting through other mechanisms. Additionally, we found that the paternal IG-DMR, previously deemed dispensable for imprinting, is required in certain tissues, demonstrating that imprinting is regulated in a tissue-specific manner. Using PRO-seq to analyze nascent transcription, we identified 30 novel transcribed regulatory elements, including 23 that are tissue-specific. These results demonstrate that different tissues have a distinctive regulatory landscape at the Dlk1-Dio3 cluster and provide insight into potential mechanisms of tissue-specific imprinting control. Together, our findings challenge the premise that Dlk1-Dio3 imprinting is regulated through a single mechanism and demonstrate that different tissues use distinct strategies to accomplish imprinted gene expression.