Engineering of translation provides an alternative regulatory layer for controlling transgene expression in addition to transcriptional regulation. Synthetic mRNA switches that modulate translation of a target gene of interest in response to an intracellular protein could be a key regulator to construct a genetic circuit. Insertion of a protein binding RNA sequence in the 5' UTR of mRNA would allow for the generation of a protein-responsive RNA switch. Here we describe the design principle of the switch and methods for tuning and analyzing its translational activity in mammalian cells.