RADICL-seq identifies general and cell type–specific principles of genome-wide RNA-chromatin interactions

Abstract

Mammalian genomes encode tens of thousands of noncoding RNAs. Most noncoding transcripts exhibit nuclear localization and several have been shown to play a role in the regulation of gene expression and chromatin remodeling. To investigate the function of such RNAs, methods to massively map the genomic interacting sites of multiple transcripts have been developed; however, these methods have some limitations. Here, we introduce RNA And DNA Interacting Complexes Ligated and sequenced (RADICL-seq), a technology that maps genome-wide RNA-chromatin interactions in intact nuclei. RADICL-seq is a proximity ligation-based methodology that reduces the bias for nascent transcription, while increasing genomic coverage and unique mapping rate efficiency compared with existing methods. RADICL-seq identifies distinct patterns of genome occupancy for different classes of transcripts as well as cell type-specific RNA-chromatin interactions, and highlights the role of transcription in the establishment of chromatin structure.

Genome-Wide Technologies to Study RNA–Chromatin Interactions

“…In 2018, ChAR-seq (chromatin-associated RNA sequencing) was reported [26]. Very recently (in 2020), RADICL-seq (RNA and DNA interacting complexes ligated and sequenced) was developed [27]. All of these methods are based on proximity ligations using a bivalent linker that can ligate to RNA at one end and to digested DNA at the other end ( Figure 2).…”

“…We discuss each of these in the following sections. [24,27,28], whereas ChAR-seq was initially tested only with Drosophila cells. GRID-seq has been tested with both Drosophila and mammalian cells.…”

“…Bonetti et al [27] found that the genomic coverage of DNA regions identified by RADICL-seq was higher than that by GRID-seq data. This could have resulted from the stronger crosslinking condition in GRID-seq and the use of AluI for DNA digestion, which introduces biases in the genome in particular for Alu elements (the enzyme is named AluI because it cleaves Alu repeats very frequently and Alu elements are located in gene-rich regions) [33].…”

“…RADICL-seq is the latest all-to-all sequencing method and was published in 2020 [27]. Despite its conceptual similarity to other all-to-all methods, the authors pointed out several key differences that improve its performance relative to other methods for studying RNA-chromatin interactions.…”

“…For RADICL-seq,~120 million reads were sequenced using the 150-cycle Illumina kit [27]. Usable reads are very limited for any of these protocols after mapping.…”