Visualizing RNA base-pairing probabilities with RNAbow diagrams

Abstract

There are many effective ways to represent a minimum free energy RNA secondary structure that make it easy to locate its helices and loops. It is a greater challenge to visualize the thermal average probabilities of all folds in a partition function sum; dot plot representations are often puzzling. Therefore, we introduce the RNAbows visualization tool for RNA base pair probabilities. RNAbows represent base pair probabilities with line thickness and shading, yielding intuitive diagrams. RNAbows aid in disentangling incompatible structures, allow comparisons between clusters of folds, highlight differences between wild-type and mutant folds, and are also rather beautiful.

FIGURE 1.

Depictions of secondary structures for the L. collosoma Spliced Leader sequence: (A) two-dimensional “airport terminal” diagram of the Minimum Free Energy (MFE) state; (B) classic “rainbow” diagram (MFE); (C) bracket notation with periods representing unpaired bases and parentheses indicating paired bases (MFE). (D) A dot plot with partition function probabilities P_ij with base i vertical and base j horizontal. Color is assigned on the basis of the logs of probabilities. (Graphics adapted from RNAstructure.) (E) ViennaRNA’s prediction (using slightly different free energy rules) with bases color-coded according to their partition function probabilities. (Graphics adapted from ViennaRNA.) (F) A Dot Plot available from ViennaRNA uses box-size proportional to probability (top triangle), but the grid obscures low-probability pairs. (G) An AllPairs RNAbow diagram with the line width and darkness proportional to the probability of the base pairs. (H) A Clusters RNAbow diagram after resolving into the two dominant clusters, with probability 0.57 (red) and 0.43 (blue); note that the MFE state (B) belongs to the less-probable blue cluster.

FIGURE 2.

The partition functions of wild-type (red) and the U22G mutant (blue) 5′ UTR of ferritin light-chain mRNA are depicted with Difference RNAbows. The colors are set proportional to the difference between the clusters such that common elements are black, while the distinct elements are either red or blue. The dramatic effect of this single nucleotide polymorphism on the secondary structure is evident. Other base changes within loop regions have less influence.

FIGURE 3.

Chemical mapping experiments indicate which bases are unpaired, and this information can also be visualized with Difference RNAbows. Here, for the 5′-UTR region of HIV-1, we compare the constrained (Schroeder et al. 2011) partition function where lowercase bases are forced to be unpaired (top, red) with the unconstrained partition function (bottom, blue).