Droplet-assisted folding of long regulatory RNAs

Abstract

Long regulatory RNA regions orchestrate complex cellular processes, including gene expression and epigenetic modifications. How these RNAs dynamically fold and refold in response to cellular signals remains poorly understood. Given that RNAs interact with ubiquitous RNA-binding proteins (RBPs) prone to form biomolecular condensates, we explore how protein droplets interacting along an RNA impact its folding process. Attached droplets prevent premature folding by competing with RNA:RNA interactions. When droplets dissolve due to cellular signals, capillary effects cause the RNA to collapse while refolding. We test this process of condensate-guided RNA folding by adapting established RNA secondary structure predictors to mimic various folding pathways and supplement this with coarse-grained simulations. We find that interactions with transient droplets robustly leads to the formation of long-range RNA contacts, which are otherwise hard to achieve. Our results compare favorably with available experimental data. We propose that this strategy, which we call droplet-assisted RNA folding, represents a previously unexplored mechanism for shaping RNA structures. Given the widespread propensity of RBPs to form condensates, this process could play a fundamental role in the structural organization, conditional reshaping, and functional regulation of long regulatory RNAs. SignificanceComplex non-coding RNA regions (e.g. lncRNAs, 3UTRs, etc.) perform several important functions in higher organisms. However, the longer an RNA, the more likely it is to misfold. How can such regulatory regions of more than 1000 nts be reliably folded, considering that premature co-transcriptional folding favors local interactions? We present a mechanism by which biomolecular condensates of RNA-binding proteins (RBPs) assist and control the folding of complex RNAs. Here, droplets of RBPs nucleate around RNA binding motifs, prevent premature misfolding through their RNA chaperone function, and allow robust folding of initially distant regions through coordinated droplet dissolution and detachment. This mechanism provides an alternative perspective on how long RNAs acquire their structures in a context that is functionally dynamic.