Four RNA families with functional transient structures

Figures & data

Figure 1. Schematic drawing for Tryptophan operon leader. The Trp codon (W) tandem is highlighted in green color in both the gene structure (the leader peptide sequence is shown) and the RNA secondary structures. The stem of the termination hairpin is colored in red in both terminator and anti-terminator structures. In the anti-terminator structure, the ribosome, stalling on the Trp tandem, impedes the formation of the first hairpin. The 5' portion (red) of the termination hairpin is then sequestered in an anti-terminator hairpin. The RNA secondary structures in Figures 1–4 are drawn via VARNA[98].

Figure 2. Schematic drawing for Levivirus (MS2). The UCS is highlighted in red color, the SD sequence is in yellow color, and the start codon is colored in green. In the left structure, the SD sequence is accessible to ribosome for translation. In the right structure, the SD pairs with the UCS, forming the LDI to impede ribosome binding.

Figure 3. Schematic drawing for HDV ribozyme. The starting point of each structure on the viral genome in relation to the self-cleavage site (this site is annotated by an orange arrow) is labeled by the corresponding number. The 5 stems of the active structure are colored using similar color scheme employed by Chadalavada et al. (2000) [12]: P1 in blue, P2 in green, P3 in yellow, P4 in dark red, P1.1 in purple. The stem P(-1) of the permissive alternative structure is colored in pink. In the inhibitory alternative structure, Alt 1, 2, and 3 disrupt the native stems of the active structure except P1 and P4.

Figure 4. Schematic drawing for SAM riboswitch. For both RNA structures, the stem of the terminator is colored in red, and the stem of the anti-anti-terminator is colored in blue. In the SAM-unbound structure, the anti-terminator is formed by the 3' portion of the anti-anti-terminator and the 5' portion of the terminator hairpin; thus, the terminator hair can no longer form. The pseudoknot is not shown in this drawing.

Table 1. The basic alignment statistics for our new alignments and the corresponding seed alignments in Rfam, if it exists. The structural quality measures allow a swift comparison between the structural annotation of our new alignments and that of the corresponding seed alignments in Rfam.

Structure	Covariation	No. BPs	Frac. Canonical BPs	No. Seqs	align. length
Trp operon leader (Rfam)	0.0653	21	0.8766	22	127
Trp operon leader (new)	0.2830	23	0.9505	29	131
HDV (Rfam-genomics)	0.0557	31	0.9821	18	115
HDV (Rfam)	0.3432	31	0.9677	33	115
HDV (new)	0.0868	30	0.996	25	155
Levivirus (new)	0.2799	48	0.9318	11	169
SAM (Rfam)	0.2980	27	0.9421	433	231
SAM (new)	0.2417	54	0.8627	85	425

For the Trp operon leader, the quality measures are calculated for the terminator structure; for HDV ribozyme, the quality measures are shown for the active structure, and provided for both the original Rfam seed alignment and the extracted genomic sequences from the seed alignment (Rfam-genomics); for the SAM riboswitch, the structure included is the SAM-bound structure. Since the levivirus is a new RNA family introduced here, only the statistics for our new Levivirus alignment is shown. Values for the covariation range from −2 to 2 and measure the relative frequency of compensatory mutations maintaining the base-pairing potential. A positive covariation implies the presence of compensatory mutations. No. BPs refers to the number of base pairs in the corresponding structure. Frac. Canonical BPs is the fraction of canonical base pairs in the alignment for the aforementioned structure. No. seqs is the number of sequences in the alignment. Align. length is the length of the gapped alignment in nucleotides. The alignment statistics are calculated using R-chie.⁹⁶

Table 2. The alignment quality measures for the transient structural features for our alignments. For Trp, the numbers refer to the anti-terminator structure.

Structure	Covariation	No. BPs	Frac. Canonical BPs
Trp operon leader (anti-terminator)	0.0635	10	0.9069
HDV (Alternative 1)	0.0437	37	0.9459
HDV (Alternative 2)	−0.0692	13	0.8677
Levivirus (transient)	0.1805	14	0.9286
SAM (SAM-unbound)	0.1449	40	0.8165

For HDV ribozyme, Alternative 1 refers to the self-cleavage-inhibitory alternative structure and Alternative 2 to the self-cleavage-permissive alternative structure. For the Levivirus alignment, transient refers to the metastable hairpins permitting the temporary translation of maturation protein. For SAM riboswitch, the structure is the SAM-unbound structure. Please see the caption of Table 1 for other definitions. The basic alignment statistics, such as the alignment size or length, are also part of Table 1 as all the alternative structures share the same alignment.

Figure 5. Arc-plot of Tryptophan operon leader made using the visualisation program R-chie [96]. The left legend specifies the percentage of canonical base-pairs in the paired alignment columns, i.e. those connected by an arc. The right legend specifies the evolutionary support (e.g., covariation, etc.) for each position in the alignment. The alignment and arcs at the top show the information for the terminator structure, whereas the bottom ones correspond to the anti-terminator structure for the same underlying alignment. The lines in each alignment correspond to the respective sequences with every box representing either a nucleotide or a gap in the respective sequence. Every arc represents a base-pair involving the respective two alignment columns. The arcs are colour-coded according to their percentage of canonical base pairs, whereas the evolutionary information supporting each base pair is encoded in the colouring of the underlying nucleotides in the base paired alignment columns, see the right legend for details. Two green blocks connected by an arc implies that this is a canonical base pair (i.e., GC, AU or GU) which corresponds to the most abundant type of base pair for this pair of alignment columns. Cyan means this is a canonical base pair but that it has an one-sided mutation with respect to the most abundant (green) base-pair. Blue refers to a canonical base-pair which differs on both sides from the most abundant (green) base-pair. Red means this is a non-canonical base pair. Unpaired nucleotides are shown in black and gaps in grey.

Figure 6. Arc-plot for the Levivirus alignment. The alignment and arcs at the top correspond to the final inhibitory structure, whereas the bottom ones correspond to the transient structure permissive for maturation protein translation. See the caption of Figure 5 for more information on arc-plots and two legends.

Figure 7. Arc-plot for the HDV ribozyme. The alignment and arcs at the top show the permissive alternative structure 2 and the active structure. The alternative structure 2, P(-1), is the 13-bp hairpin on the left side on top of the arc diagram. The bottom arcs correspond to the inhibitory alternative structure 1. See the caption of Figure 5 for more information on arc-plots and two legends.

Figure 8. Arc-plot of SAM riboswitch. The top covariation and arcs correspond to the SAM-bound conformation and the bottom ones refer to the SAM-unbound conformation.

Darty, K., Denise, A., & Ponty, Y. VARNA: Interactive drawing and editing of the RNA secondary structure. Bioinformatics 2009; 25 (15):1974-1975.

 PubMed Web of Science ®Google Scholar

Chadalavada, D. M., Knudsen, S. M., Nakano, S. & Bevilacqua, P. C. A role for upstream RNA structure in facilitating the catalytic fold of the genomic hepatitis delta virus ribozyme. J Mol Biol 2000; 301: 349-367.

 PubMed Web of Science ®Google Scholar

Lai, D., Proctor, J., Zhu, J. & Meyer, I. R-chie: a web server and r package for visualizing RNA secondary structures. Nucleic Acids Res 2012; 40:e95.

 PubMed Web of Science ®Google Scholar