Translational regulation by mRNA secondary structures in yeast
Translation in Saccharomyces cerevisiae can be regulated by secondary structures within an mRNA, controlling both the level, and identity, of the encoded protein. In this study RNA aptamers specific for theophylline and the dye Hoechst 33342 were used to replace the hairpin structure within the HIV-1 frameshift signal. The theophylline aptamer RNA structure was a weak stimulator of frameshifting, but increased frameshifting in response to the ligand was not observed. Aptamers were also placed within the 5’UTR of a reporter mRNA to examine whether ligand-dependent inhibition of scanning initiation could be demonstrated. Within the 5’ UTR, a single copy of the Hoechst 33342, kanamycin, tobramycin or theophylline aptamer reduced the level of reporter activity. However only the theophylline aptamer was shown to inhibit translation in a ligand-dependent manner. This effect could be slightly enhanced when theophylline aptamers were placed in tandem, and when the temperature of S. cerevisiae growth was reduced from 30°C to 12° C. Overall, the results indicate that in vivo, RNA aptamers cannot easily be used to regulate gene expression in a ligand-dependent manner. Secondly, the role of mRNA secondary structure in regulating readthrough of a retrotransposon stop codon was investigated. Expression of the pol ORF in the Candida albicans retrotransposon Tca2 is prevented by a stop codon at the end of the gag ORF. It has been suggested that pol expression occurs via readthrough of this stop codon, possibly regulated by a pseudoknot downstream of the stop codon. This study has shown that Tca2 is widespread in C. albicans.