Orthogonal Protein-Responsive mRNA Switches for Mammalian Synthetic Biology

The lack of available genetic modules is a fundamental issue in mammalian synthetic biology. Especially, the variety of genetic parts for translational control are limited. Here we report a new set of synthetic mRNA-based translational switches by engineering RNA-binding proteins (RBPs) and RBP-binding RNA motifs (aptamers) that perform strong translational repression. We redesigned the RNA motifs with RNA scaffolds and improved the efficiency of the repression to target RBPs. Using new and previously reported mRNA switches, we demonstrated that the orthogonality of translational regulation was ensured among five different RBP-responsive switches. Moreover, the new switches functioned not only with plasmid introduction, but also with RNA-only delivery, which provides a transient and safer regulation of expression. The translational regulators using RNA-protein interactions provide an alternative strategy to construct complex genetic circuits for future cell engineering and therapeutics.