Stable G-quadruplex enabling sequences are selected against by the context-dependent codon bias

The distributions of secondary structural elements appear to differ between coding regions (CDS) of mRNAs compared to the untranslated regions (UTRs), presumably as a mechanism to fine-tune gene expression, including efficiency of translation. However, a systematic and comprehensive analysis of secondary structure avoidance because of potential bias in codon usage is difficult as some of the common secondary structures, such as, hairpins can be formed by numerous sequence combinations. Using G-quadruplex (GQ) as the model secondary structure we studied the impact of codon bias on GQs within the CDS. Because GQs can be predicted using specific consensus sequence motifs, they provide an excellent platform for investigation of the selectivity of such putative structures at the codon level. Using a bioinformatics approach, we calculated the frequencies of putative GQs within the CDS of a variety of species. Our results suggest that the most stable GQs appear to be significantly underrepresented within the CDS, through the use of specific synonymous codon combinations. Furthermore, we identified many peptide sequence motifs in which silent mutations can potentially alter translation via stable GQ formation. This work not only provides a comprehensive analysis on how stable secondary structures appear to be avoided within the CDS of mRNA, but also broadens the current understanding of synonymous codon usage as they relate to the structure-function relationship of RNA.