Conformation and structural dynamics of the Xist lncRNA A-repeats

LncRNAs are emerging to play crucial roles in the regulation of many essential cellular processes and have been linked to human disease, but a detailed understanding of their structure and how this relates to underlying molecular mechanisms is still limited. The structure that a lncRNA adopts can interconvert between multiple conformations. However, characterizing the structure and dynamics is challenging given their large size. Here, we present an integrated approach, combining biochemical and biophysical techniques to investigate the core structural elements and conformational dynamics of the A-repeats of the lncRNA Xist. We combine chemical RNA structure probing, SAXS, NMR-spectroscopy and cryo-EM to comprehensively describe the conformational landscape of the Xist A-repeats. We show that under native-like conditions, the A-repeats are modular, comprising building blocks made from stable AUCG tetraloop hairpins and inter-repeat dimers separated by flexible uracil-rich regions. The structural core of the A-repeats involves dimerization of sequential repeats to form two subdomains, comprising repeats 1-4 and 5-8. The overall topology of the A-repeats is dynamic, with structural variability linked to the uracil-rich linker regions. Our results rationalize context and buffer-dependent structural variations of the Xist lncRNA. The integrative approach presented here establishes a general pipeline for investigating lncRNA structure and dynamics. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=98 SRC="FIGDIR/small/501616v2_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1761cbeorg.highwire.dtl.DTLVardef@15905e0org.highwire.dtl.DTLVardef@190bf0corg.highwire.dtl.DTLVardef@1eeca5a_HPS_FORMAT_FIGEXP M_FIG C_FIG