Sequencing data to detect RNA-small molecule interactions

RNA structure formation is significantly influenced by single nucleotide polymorphisms (SNPs) and somatic mutations. Consequently, RNA structural modulation induced by genetic variants can alter binding affinity to RNA-targeting small-molecule drugs, potentially affecting their therapeutic efficacy significantly. Thus, for the advancement of RNA-targeting small-molecule drug discovery, it is crucial to comprehensively identify variant-specific RNA-small-molecule interactions and identify genetic mutations that alter binding affinity. On the other hand, conventional studies of RNA-small-molecule interactions have, due to the technical limitations of existing large-scale detection technologies, evaluated binding only against the reference genome, thereby ignoring the impact of genetic variants. We introduce BIVID-MaP (Binding- and Vinyl-Quinazolinone-Induced Deletion-Based Mutational Profiling), the method for identifying variant-specific RNA-small-molecule interactions. BIVID-MaP detects small-molecule-binding sites by combining a binding-dependent covalent modification reaction with reverse transcription deletion profiling via deep sequencing. The reverse transcription deletions detected by this method are mostly single nucleotides, and the sequence reads contain nearly full-length sequence information, excluding the deleted base. This feature enables the differentiation of reads for each variant and the individual detection of deletions. By providing novel insights into RNA structure-small-molecule interactions specific to individual genetic variants, this study is expected to contribute as a foundational technology advancing both RNA structural biology and RNA-targeting drug discovery.