Genome-wide colocalization of RNA–DNA interactions and fusion RNA pairs

Abstract

Fusion transcripts are used as biomarkers in companion diagnoses. Although more than 15,000 fusion RNAs have been identified from diverse cancer types, few common features have been reported. Here, we compared 16,410 fusion transcripts detected in cancer (from a published cohort of 9,966 tumor samples of 33 cancer types) with genome-wide RNA–DNA interactions mapped in two normal, noncancerous cell types [using iMARGI, an enhanced version of the mapping of RNA–genome interactions (MARGI) assay]. Among the top 10 most significant RNA–DNA interactions in normal cells, 5 colocalized with the gene pairs that formed fusion RNAs in cancer. Furthermore, throughout the genome, the frequency of a gene pair to exhibit RNA–DNA interactions is positively correlated with the probability of this gene pair to present documented fusion transcripts in cancer. To test whether RNA–DNA interactions in normal cells are predictive of fusion RNAs, we analyzed these in a validation cohort of 96 lung cancer samples using RNA sequencing (RNA-seq). Thirty-seven of 42 fusion transcripts in the validation cohort were found to exhibit RNA–DNA interactions in normal cells. Finally, by combining RNA-seq, single-molecule RNA FISH, and DNA FISH, we detected a cancer sample with EML4-ALK fusion RNA without forming the EML4-ALK fusion gene. Collectively, these data suggest an RNA-poise model, where spatial proximity of RNA and DNA could poise for the creation of fusion transcripts.

RNA promotes the formation of spatial compartments in the nucleus

“…Moreover, we recently showed that proximityligation methods can fail to identify pairwise contacts between molecules that are organized within nuclear compartments because these methods only identify interactions where components are close enough in space to be directly ligated 54 . Consistent with this observation, existing RNA-DNA proximity-ligation methods fail to identify known RNA-DNA contacts that are contained within various well-established nuclear bodies, such as nucleoli, histone locus bodies (HLBs), and Cajal bodies [50][51][52][53] .…”

“…However, this approach is limited to examining a small number of simultaneous interactions and therefore requires a priori knowledge of which RNAs and nuclear structures to explore. An alternative approach is genomic mapping of RNA-DNA contacts using proximity-ligation methods [49][50][51][52][53] . While these approaches can provide genome-wide pairwise maps of RNA-DNA interactions, they do not provide information about the 3D organization of these molecules in the nucleus.…”

Stress-induced RNA–chromatin interactions promote endothelial dysfunction

“…Recent technological developments have made it possible to assay DNA-DNA and RNA-chromatin interactions in situ in a genome-wide manner [6][7][8][9][10][11] . Among these tools, in situ mapping of RNA-genome interactome (iMARGI) enables all-RNA-versusthe-genome analyses that can simultaneously identify many caRNAs and their respective genomic interaction loci 7,8 . This feature helped to reveal a large number of caRNAs, including those attached to other chromosomes 7,12 .…”

Dynamic changes in RNA-chromatin interactome promote endothelial dysfunction

“…In situ MARGI was performed as described in our recent report 5 . Briefly, iMARGI started with crosslinking cells using 1% formaldehyde, collecting nuclei, followed by fragmenting RNA and DNA in nuclei using RNase I and restriction enzyme AluI.…”

“…Among these efforts, we have developed in situ mapping of RNA-genome interaction (iMARGI) assay which uses a bivalent linker to ligate to nuclear RNA and DNA followed by circularization for library construction 4,5 . By utilizing the advantages of iMARGI, we observed that caRNAs are not only associated with the genomic sequences at or near their transcription sites, but are also attached to distal genomic sequences on the same or other chromosomes.…”