Cell 165, 1267–1279 (2016)

Credit: ZHIPENG LU AND HOWARD Y CHANG

Base-paired duplexes are the fundamental unit of RNA structure and RNA–RNA interactions. Chemical probing and informatics tools that define paired and unpaired regions have enabled RNA structural analyses within cells, but these approaches generally uncover only short-range interactions and simple RNA motifs. Lu et al. now report PARIS (psoralen analysis of RNA interactions and structures) as a method for mapping RNA duplexes on a transcriptome-wide scale. The approach relies on UV-mediated psoralen crosslinking of duplex regions in cells followed by analysis of the isolated crosslinked fragments by reverse transcription and next-generation DNA sequencing (NGS). 'Duplex groups' are then assigned by informatics analysis to infer candidate intramolecular stem-loop structures or in trans RNA–RNA interactions within cells. PARIS applied to several human and mouse cell lines revealed that duplex formation between regions separated by hundreds or thousands of nucleotides is widespread in the transcriptome. Further, the cross-species comparisons and multiple NGS reads per locus identified more complex structures (such as pseudoknots) and abundant alternative RNA conformations between duplex groups. Application of PARIS with other probing methods yielded a detailed duplex map of Xist, providing structural evidence that A-repeat assembly in this lncRNA is essential for binding of SPEN, a factor involved in X-chromosome inactivation. PARIS adds to the RNA toolbox for mapping RNA structure within cells and exploring its functional roles.