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Article

Analysis of the H19ICR Insulator

, , , , &
Pages 3499-3510 | Received 20 Nov 2006, Accepted 15 Feb 2007, Published online: 27 Mar 2023
 

Abstract

Transcriptional insulators are specialized cis-acting elements that protect promoters from inappropriate activation by distal enhancers. The H19 imprinting control region (ICR) functions as a CTCF-dependent, methylation-sensitive transcriptional insulator. We analyzed several insertional mutations and demonstrate that the ICR can function as a methylation-regulated maternal chromosome-specific insulator in novel chromosomal contexts. We used chromosome conformation capture and chromatin immunoprecipitation assays to investigate the configuration of cis-acting elements at these several insertion sites. By comparing maternal and paternal organizations on wild-type and mutant chromosomes, we hoped to identify mechanisms for ICR insulator function. We found that promoter and enhancer elements invariably associate to form DNA loop domains at transcriptionally active loci. Conversely, active insulators always prevent these promoter-enhancer interactions. Instead, the ICR insulator forms novel loop domains by associating with the blocked promoters and enhancers. We propose that these associations are fundamental to insulator function.

We thank Judy Kassis, Michael Miller, and Claudia Gebert for helpful discussions and advice.

This work was funded by the NICHD Intramural Research Program.

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