Positive selection acting on splicing motifs reflects compensatory evolution

Abstract

We have used comparative genomics to characterize the evolutionary behavior of predicted splicing regulatory motifs. Using base substitution rates in intronic regions as a calibrator for neutral change, we found a strong avoidance of synonymous substitutions that disrupt predicted exonic splicing enhancers or create predicted exonic splicing silencers. These results attest to the functionality of the hexameric motif set used and suggest that they are subject to purifying selection. We also found that synonymous substitutions in constitutive exons tend to create exonic splicing enhancers and to disrupt exonic splicing silencers, implying positive selection for these splicing promoting events. We present evidence that this positive selection is the result of splicing-positive events compensating for splicing-negative events as well as for mutations that weaken splice-site sequences. Such compensatory events include nonsynonymous mutations, synonymous mutations, and mutations at splice sites. Compensation was also seen from the fact that orthologous exons tend to maintain the same number of predicted splicing motifs. Our data fit a splicing compensation model of exon evolution, in which selection for splicing-positive mutations takes place to counter the effect of an ongoing splicing-negative mutational process, with the exon as a whole being conserved as a unit of splicing. In the course of this analysis, we observed that synonymous positions in general are conserved relative to intronic sequences, suggesting that messenger RNA molecules are rich in sequence information for functions beyond protein coding and splicing.[Supplemental material is available online at www.genome.org.]In addition to sequences that specify their cognate polypeptides, the open reading frames of eukaryotic messenger RNAs (mRNAs) are likely to contain additional information governing the functioning of these molecules; these functions include exon splicing and mRNA transport, localization, stability, and translation. If so, the evolution of mRNA sequence would be constrained by purifying selection against the loss of this information. This selection should be evident by examining the rate and quality of base substitutions that do not alter protein coding (synonymous substitutions) but may alter one of the abovementioned functions.Mutation rates in open reading frames have been extensively characterized by two parameters: K a , the rate of mutations that result in amino acid substitutions; and K s , the rate at synonymous sites. The latter has most often been assumed to reflect neutral change and has been used to normalize the mutation rates in a given gene, with the K a /K s ratio serving as an inverse measure of protein sequence conservation (Hurst 2002). More recently, the neutrality of K s has been challenged, with accumulating evidence for selection acting at synonymous sites (for reviews, see Chamary et al. 2006;Xing and Lee 2006). Some of this evidence has been based on comparisons between mutation rates in predicted exonic splicing e...

The Silent Sway of Splicing by Synonymous Substitutions

“…This result reproduces the finding that the A to C mutation at position 7 rescues the C to T mutation at position 6 either through inhibiting the binding of Sam68, or by preventing the binding of another inhibitory protein (33,34). Similar examples of synergistic or compensatory mutations are observed to occur at the Tra2-␤1 (SRSF10) binding site (positions 18 -28), at the 3Ј inhibitory secondary structure (positions [33][34][35][36][37][38][39][40][41][42], and at the 5Ј splice site (21, 23, 32, 35) (Figs. 2, 3, and 5, Table 1).…”

“…The presence of splicing regulatory elements at many of these conserved synonymous positions further suggests that there has been positive selection throughout evolution for specific splicing events. Positive selection due to splicing within splicing regulatory elements and around intron/exon junctions has been previously suggested and documented, but only recently experimentally analyzed by testing for missense and nonsense SNPs associated with disease (4,7,(37)(38)(39). However, positive selection is also at work in the genetic code, masking changes that may have occurred due to splicing.…”

RNA landscape of evolution for optimal exon and intron discrimination

“…A direct consequence of the degeneracy of the binding motifs is that SREs are highly ubiquitous. Therefore, the higher-order constraints are also reflected in the base composition, because exonic (intronic) nucleotide substitutions toward EIEs (IIEs) are favored for the discrimination between exons and introns (29). However, we could not distinguish whether exonic and intronic sequences adapted to the specificity of the splicing machinery during early evolution or vice versa.…”

Unexpected selection to retain high GC content and splicing enhancers within exons of multiexonic lncRNA loci

“…Additionally, similar to our conclusions of a greater conservation of ESEs close to lncRNA exon boundaries, the authors also showed that the selective pressure on ESEs was also stronger close to splice sites. Several subsequent analyses of ESEs within protein-coding sequences reached similar conclusions either using multispecies comparisons or larger SNV data sets in human Ke et al 2008;Cáceres and Hurst 2013).…”