Translation regulation by a Guanidine-II riboswitch

Comprehensive mutational analysis of transcriptional riboswitches has been used to probe the energetic intricacies of interplay between the aptamer and expression platform, but translational riboswitches have been inaccessible to massively parallel techniques. The Guanidine-II riboswitch is an exclusively translational class. We have integrated RelE cleavage with next generation sequencing to quantify ligand-dependent changes in translation initiation for all single and double mutations of the Pseudomonas aeruginosa Gdm-II riboswitch, a total of more than 23000 variants. This comprehensive examination of a translational riboswitch's expression platform illuminates how the riboswitch is precisely tuned and tunable with regard to ligand sensitivity, amplitude of expression between ON and OFF states, and cooperativity of ligand binding.