Active enhancers strengthen insulation by RNA-mediated CTCF binding at chromatin domain boundaries

• Type of Material: Article
• Authors: Zubairul Islam, National Centre for Biological Sciences; Bharath Saravanan, National Centre for Biological Sciences; Kaivalya Walavalkar, National Centre for Biological Sciences; Umer Farooq, National Centre for Biological Sciences; Anurag Kumar Singh, National Centre for Biological Sciences; Radhakrishnan Sabarinathan, National Centre for Biological Sciences; Jitendra Thakur, Emory University; Awadhesh Pandit, National Centre for Biological Sciences; Steven Henikoff, Fred Hutchinson Cancer Research Center; Dimple Notani, National Centre for Biological Sciences
• Language: English
• Date: 2023-01-01
• Publisher: Cold Spring Harbor Laboratory Press
• Publication Version: Final Published Version
• Copyright Statement: © 2023 Islam et al.; Published by Cold Spring Harbor Laboratory Press
• License: Creative Commons Attribution-NonCommercial 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1101/gr.276643.122
• Title of Journal or Parent Work: Genome Research
• Volume: 33
• Issue: 1
• Start Page: 1
• End Page: 17
• Keywords: Enhancer Elements, Genetic; CCCTC-Binding Factor; Promoter Regions, Genetic; RNA; Binding Sites; Chromatin
• Research Categories: Health Sciences, Oncology

Abstract

Vertebrate genomes are partitioned into chromatin domains or topologically associating domains (TADs), which are typically bound by head-to-head pairs of CTCF binding sites. Transcription at domain boundaries correlates with better insulation; however, it is not known whether the boundary transcripts themselves contribute to boundary function. Here we characterize boundary-associated RNAs genome-wide, focusing on the disease-relevant INK4a/ARF and MYC TAD. Using CTCF site deletions and boundary-associated RNA knockdowns, we observe that boundary-associated RNAs facilitate recruitment and clustering of CTCF at TAD borders. The resulting CTCF enrichment enhances TAD insulation, enhancer–promoter interactions, and TAD gene expression. Importantly, knockdown of boundary-associated RNAs results in loss of boundary insulation function. Using enhancer deletions and CRISPRi of promoters, we show that active TAD enhancers, but not promoters, induce boundary-associated RNA transcription, thus defining a novel class of regulatory enhancer RNAs.