Structural and functional characterization of ribosomal protein gene introns in sponges

Abstract

Ribosomal protein genes (RPGs) are a powerful tool for studying intron evolution. They exist in all three domains of life and are much conserved. Accumulating genomic data suggest that RPG introns in many organisms abound with non-protein-coding-RNAs (ncRNAs). These ancient ncRNAs are small nucleolar RNAs (snoRNAs) essential for ribosome assembly. They are also mobile genetic elements and therefore probably important in diversification and enrichment of transcriptomes through various mechanisms such as intron/exon gain/loss. snoRNAs in basal metazoans are poorly characterized. We examined 449 RPG introns, in total, from four demosponges: Amphimedon queenslandica, Suberites domuncula, Suberites ficus and Suberites pagurorum and showed that RPG introns from A. queenslandica share position conservancy and some structural similarity with "higher" metazoans. Moreover, our study indicates that mobile element insertions play an important role in the evolution of their size. In four sponges 51 snoRNAs were identified. The analysis showed discrepancies between the snoRNA pools of orthologous RPG introns between S. domuncula and A. queenslandica. Furthermore, these two sponges show as much conservancy of RPG intron positions between each other as between themselves and human. Sponges from the Suberites genus show consistency in RPG intron position conservation. However, significant differences in some of the orthologous RPG introns of closely related sponges were observed. This indicates that RPG introns are dynamic even on these shorter evolutionary time scales.

Figure 1. Size distribution of the RPG introns in sponge A. queenslandica (black bars), cnidarian N. vectensis (gray bars), and placozoan T. adhaerens (white bars).

Figure 2. Over-represented elements in sponge A. queenslandica RPG introns composed of three motifs.

Full tripartite motifs are circled. The E-value and combined p-value were extracted from MEME.

Figure 3. Combinatorial grouping of 10 species according to the number of shared introns

. Gray squares indicate presence of introns.

Figure 4. C/D box snoRNAs conserved in sponges and human.

(A) Intron-mapping of RPS12 genes from representative species. White triangles indicate positions of the introns and gray triangles indicate presence of SNORD100. The number in the triangle denotes the intron phase and the number in brackets intron length. The thin line indicates the 5′ UTR region. (B) The target site of SNORD100 in 18S rRNA is marked with an asterisk. (C) Alignment of SNORD100 orthologs from various metazoans. (D) Alignment of SNORD24 orthologs. One methylation guide site in S. domuncula, S. ficus and S. pagurorum is not conserved. (E) Target sites of SNORD24 are conserved in sponge and marked with dot and asterisk. (F) Orthologs of SNORD83 with conserved methylation (Me) guide site of unknown target.

Figure 5. H/ACA snoRNAs conserved in the fifth and sixth introns (I5, I6) of the RPL13A gene in S. domuncula (SD), S. ficus (SF) and S. pagurorum (SP) and in the sixth intron of the same gene in A. queenslandica (AQ).

All essential snoRNA elements are conserved and a putative pseudouridylation (PU) guide site is designated.

Figure 6. A potential 28S rRNA target (A) and secondary structure of a novel snoRNA found in the last intron of the RPS19 gene in S. domuncula (B), S. ficus (C) and S. pagurorum (D).

Figure 7. Multiple alignment of selected introns from three species of the genus Suberites: S. domuncula (SD), S. ficus (SF) and S. pagurorum (SP).

A large insertion is shown in the fourth intron of the SF RPL5 gene (A), changes in first introns of RPS15A gene (B), smaller changes between fifth introns of the RPS18 gene (C) and relatively high conservation in the sixth intron of RPP0 gene (D).