Genome-Wide Analysis of Stable RNA Secondary Structures across Multiple Organisms Using Chemical Probing Data: Insights into Short Structural Motifs and RNA-Targeting Therapeutics

Small molecules targeting specific RNA-binding sites, including stable and transient RNA structures, are emerging as effective pharmacological approaches for modulating gene expression. However, little is understood about how stable RNA secondary structures are shared across organisms, which is an important factor in controlling drug selectivity. In this study, I provide an analytical pipeline named RNA secondary structure finder (R2S-Finder) to discover short, stable RNA structural motifs in humans, <i>Escherichia coli</i> (<i>E. coli</i>), SARS-CoV-2, and Zika virus by leveraging existing in vivo and in vitro genome-wide chemical RNA-probing datasets. I found several common features across the organisms. For example, apart from the well-documented tetraloops, AU-rich tetraloops are widely present in different organisms. I also validated that the 5' untranslated region (UTR) contains a higher proportion of stable structures than the coding sequences in humans and Zika virus. In general, stable structures predicted from in vitro (protein-free) and in vivo datasets are consistent across different organisms, indicating that stable structure formation is mostly driven by RNA folding, while a larger variation was found between in vitro and in vivo data for certain RNA types, such as human long intergenic noncoding RNAs (lincRNAs). Finally, I predicted stable three- and four-way RNA junctions that exist under both in vivo and in vitro conditions and can potentially serve as drug targets. All results of stable structures, stem-loops, internal loops, bulges, and <i>n</i>-way junctions have been collated in the R2S-Finder database (https://github.com/JingxinWangLab/R2S-Finder), which is coded in hyperlinked HTML pages and tabulated in CSV files.