Active enhancers strengthen insulation by RNA-mediated CTCF binding at chromatin domain boundaries
- Zubairul Islam1,2,6,
- Bharath Saravanan1,2,6,
- Kaivalya Walavalkar1,
- Umer Farooq1,3,
- Anurag Kumar Singh1,
- Radhakrishnan Sabarinathan1,
- Jitendra Thakur4,
- Awadhesh Pandit1,
- Steven Henikoff5 and
- Dimple Notani1
- 1National Center for Biological Sciences, Tata Institute for Fundamental Research, Bangalore, Karnataka 560065, India;
- 2Sastra Deemed University, Thanjavur, Tamil Nadu 613401, India;
- 3The University of Trans-Disciplinary Health Sciences and Technology, Bangalore, Karnataka 560064, India;
- 4Department of Biology, Emory University, Atlanta, Georgia 30322, USA;
- 5Basic Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, Washington 98109, USA
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↵6 These authors contributed equally to this work.
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.
Footnotes
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[Supplemental material is available for this article.]
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Article published online before print. Article, supplemental material, and publication date are at https://www.genome.org/cgi/doi/10.1101/gr.276643.122.
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Freely available online through the Genome Research Open Access option.
- Received January 26, 2022.
- Accepted November 10, 2022.
This article, published in Genome Research, is available under a Creative Commons License (Attribution-NonCommercial 4.0 International), as described at http://creativecommons.org/licenses/by-nc/4.0/.
