oRNAment: a database of putative RNA binding protein target sites in the transcriptomes of model species

Abstract

Protein-RNA interactions are essential for controlling most aspects of RNA metabolism, including synthesis, processing, trafficking, stability and degradation. In vitro selection methods, such as RNAcompete and RNA Bind-n-Seq, have defined the consensus target motifs of hundreds of RNA-binding proteins (RBPs). However, readily available information about the distribution features of these motifs across full transcriptomes was hitherto lacking. Here, we introduce oRNAment (o RNA motifs enrichment in transcriptomes), a database that catalogues the putative motif instances of 223 RBPs, encompassing 453 motifs, in a transcriptome-wide fashion. The database covers 525 718 complete coding and non-coding RNA species across the transcriptomes of human and four prominent model organisms: Caenorhabditis elegans, Danio rerio, Drosophila melanogaster and Mus musculus. The unique features of oRNAment include: (i) hosting of the most comprehensive mapping of RBP motif instances to date, with 421 133 612 putative binding sites described across five species; (ii) options for the user to filter the data according to a specific threshold; (iii) a user-friendly interface and efficient back-end allowing the rapid querying of the data through multiple angles (i.e. transcript, RBP, or sequence attributes) and (iv) generation of several interactive data visualization charts describing the results of user queries. oRNAment is freely available at http://rnabiology.ircm.qc.ca/oRNAment/.

Figure 1.

The oRNAment database contains 453 motifs attributable to 223 RBPs in 5 species. (i) Motifs obtained for each RBP come from RNAcompete (red segment) and RBNS (dark grey segment) experiments. Links shown between RBPs (light grey lines) denote those that were assessed by both methods. Coloured dots show the species-specificity of each motif according to Ray et al. (12). There are 181 RBPs with binding specificities in the species included in the database and 42 from external species. (ii) Upset plot showing the distribution of interrogated species-specific RBPs across all five species.

Figure 2.

The oRNAment computational pipeline. (i) For a given transcript, the algorithm linearly scans for subsequences of length 7 and (ii) reports only those that have an MSS higher than the threshold, represented by the dashed line (table look-up, exemplified by the arrows, only shown for the second and fourth sequentially scanned 7-mers; sum of MSS’ used as denominator for MSS’% computation in bold). (iii) oRNAment reports all motif instances in all transcripts across five species. (iv) oRNAment reasonably predicts RBP binding sites observed by eCLIP in human K562 and HepG2 cells (blue bars in histogram), as shown for the five motifs bound by the HNRNPK RBP, in comparison to the same number of random sequences (orange bars).

Figure 3.

Examples of the figures generated by oRNAment when searching for motifs in specific RNAs or RBPs. Upon a user's query, either by transcript (i–v) or by RBP (vi-ix), multiple figures summarizing the results are provided. (i–v) When searching by transcripts (here the cen mRNA in Drosophila), oRNAment provides: (i) a treemap of the most abundant RBP motif instances (likewise shown when searching by attributes); or (ii) a histogram of the same results; (iii) a polar plot showing in which subregion of the transcript RBP motif instances are observed (here in cen); or (iv) a histogram of the same results; (v) a box plot of the distribution of RBP motif instances in all transcripts queried (here, the boxplot shows the distribution of the number of motif instances among the two isoforms of cen). (vi–ix) When searching by RBP, oRNAment provides: (vi) a doughnut plot showing in which gene biotypes putative binding sites for the queried RBP are observed (here for SRSF9); or (vii) a histogram of the same results; and (viii) a radar plot showing in what transcript subregion putative binding sites for the queried RBP are observed; or (ix) a histogram of the same results. All search functionalities provide a table from which the user can access gene-level or transcript-level details. (x–xi) By selecting a gene/transcript and RBP pair, oRNAment will provide: (x) a scatter plot showing the position of each putative RBP binding site and corresponding MSS scores, here above the 50% MSS’ threshold respectively for each motif of the shep RBP on the cen mRNA. The transcript positions, on the x-axis, end at the last motif instance + 10 nucleotides; and (xi) a predicted 2D structure of the cen transcript as established by RNAfold with default parameters.

Figure 4.

Combined visualization of putative binding sites for three RBPs in two genes through a standard scatter plot and an embedded Integrative Genome Browser. (i) Example of oRNAment transcript-level view scatter plot of three RBPs (ANKHD1, FUS and lark) for two mRNAs (SMAD2 and SMAD4) and (ii) Integrative Genome Browser view incorporating the same results when searching for their loci [IGV Locus search input in the form: 18:47783250–47964001 18:51000898–51113761 (i.e. with a space separating the coordinates)]. Two examples of corresponding motif instances (lark in SMAD2 and FUS in SMAD4) between the two types of analysis are shown.