Strong promoters are mutationally robust

Mutational robustness is the persistence of a phenotype upon mutation. It facilitates molecular evolution and has been characterized in a variety of biological systems, but studies of prokaryotic promoters are limited. Prokaryotic promoters are non-coding DNA sequences that regulate transcription. The main housekeeping promoters (σ70) share two sequence motifs called the -35 and the -10 box that are spaced 17±1 base pairs (bps) apart. The sequence of these boxes, their distance, and the existence of multiple nearby boxes can determine promoter strength, the ability of a promoter to drive high levels of transcription. Here we used computational modelling to show that the mutational robustness of σ70 promoters – the persistence of promoter strength upon mutation – correlates with promoter strength itself. Mutational robustness is also influenced by potential overlaps between -10 and -35 boxes, and by specific positions within the boxes. It is higher when the boxes are exactly 17 bps apart. These findings can be partially explained by the flexibility of -35 boxes, which enables adjacent bases to create overlapping and new -35 boxes. Our work can help to engineer synthetic promoters for strength and robustness, and to understand the dynamics of promoter evolution.