R2DT: a comprehensive platform for visualizing RNA secondary structure

Abstract

RNA secondary (2D) structure visualization is an essential tool for understanding RNA function. R2DT is a software package designed to visualize RNA 2D structures in consistent, recognizable, and reproducible layouts. The latest release, R2DT 2.0, introduces multiple significant features, including the ability to display position-specific information, such as single nucleotide polymorphisms or SHAPE reactivities. It also offers a new template-free mode allowing visualization of RNAs without pre-existing templates, alongside a constrained folding mode and support for animated visualizations. Users can interactively modify R2DT diagrams, either manually or using natural language prompts, to generate new templates or create publication-quality images. Additionally, R2DT features faster performance, an expanded template library, and a growing collection of compatible tools and utilities. Already integrated into multiple biological databases, R2DT has evolved into a comprehensive platform for RNA 2D visualization, accessible at https://r2dt.bio.

Graphical Abstract

Figure 1.

(A) A fragment of a rabbit small ribosomal subunit rRNA sequence visualized using the human template with Infernal posterior probabilities reflecting alignment confidence shown as coloured circles (RNAcentral ID URS000086855E_9986). The highlighted regions help identify species-specific differences between the human and the rabbit rRNAs. (B) Normalized DMS reactivities for SLs SL1–SL4 of SARS-CoV-2 (data from [21]). (C) Cripavirus IRES with three pseudoknots (RNAcentral ID URS0000A7638F_12 136). (D) Bridge RNA from Escherichia coli (INSDC accession CP147105.1/771271–771455) visualized using the new template-free mode (data from [22]).

Figure 2.

(A) Screenshot of the RNAcanvas user interface showing human mitochondrially encoded 12S rRNA (MT-RNR1). The layout and the 2D structure were generated by R2DT, while the fonts, colour, background, and non-canonical interactions were configured in RNAcanvas. (B) Screenshot of the XRNA-React website showing a 5S rRNA.

Figure 3.

Example diagrams illustrating different constrained folding modes in R2DT. The 5S sequence from Halobacteroides halobius is visualized using a 5S template from Bacillus subtilis (INSDC accession CP003359.1/20186–20360). (A) Default R2DT output without constrained folding. (B) Local folding mode where RNAfold is used to predict the structure for the insertion relative to the template. (C) Global folding mode with additional base pairs predicted throughout the structure. (D) An example of using global folding mode with an exclusion mask. In this case all unpaired nucleotides aligned to the template were prevented from forming base pairs. Predicted base pairs are shown as dashed lines. Changes relative to default output are highlighted in blue boxes.

Figure 4.

Example R2DT templates based on consensus 2D structures of Rfam families visualized using R2R and RNArtist in the left and right subpanels, respectively. (A) Cobalamin riboswitch (Rfam ID RF00174). (B) FMN riboswitch (Rfam ID RF00050). The R2R layouts contain major overlaps between the structural elements making it difficult to see the nucleotides, while the RNArtist layouts prevent the overlaps and are more legible. For both families, R2DT 2.0 uses the RNArtist layouts by default.

Figure 5.

(A) Screenshot of the 2D and 3D components for 23S rRNA from E. coli, in PDB entry 3CC2. (B) Screenshot of the 2D and 3D components for 16S rRNA from E. coli, in PDB entry 5J7L, showing a kink-turn RNA 3D module and its non-canonical base pairs annotated using FR3D Python according to the Leontis–Westhof nomenclature [25]. The circles, squares, and triangles represent the Watson–Crick, Hoogsteen, and sugar edges of the nucleotides, respectively. In panels (A) and (B), nucleotides G691 and G705 are selected in both the 2D and 3D views. These nucleotides are highlighted and marked with black arrows on the left side of each panel.