Structural insights into higher-order natural RNA-only multimers

RNA-only complexes adopt intricate three-dimensional structures to fulfill diverse functions independently of protein partners. Although multimeric RNA-only structures have been engineered in synthetic RNA nanomaterials, naturally occurring RNA-only complexes have primarily been observed as monomers or dimers, leaving higher-order assemblies largely unexplored. ROOL (rumen-originating, ornate, large) and GOLLD (giant, ornate, lake- and Lactobacillales-derived) RNAs are conserved non-coding RNAs with complex secondary structures, but their high-resolution architectures remain unknown. Here, we determine the cryo-electron microscopy structures of UCC118-Rool RNA, Sag-Golld RNA and Env38-Golld RNA at 1.96-2.98 Å resolution, revealing their distinct hexameric, decameric and tetradecameric assemblies. These higher-order architectures are stabilized by an array of tertiary motifs such as kissing loops and tetraloop-receptor motifs, underscoring the conserved principles of RNA self-assembly. By elucidating the molecular details of these higher-order RNA-only assemblies, this study expands our understanding of RNA-based architectures and broadens the scope of RNA structural biology.