Abstact

Synthetic RNA devices are engineered to control gene expression and offer great potential in both biotechnology and clinical applications. Here, we present multidisciplinary structural and biochemical data for a tetracycline (Tc)-responsive RNA device (D43) in both ligand-free and bound states, providing a structure-dynamical basis for signal transmission. Activation of self-cleavage is achieved via ligand-induced conformational and dynamical changes that stabilize the elongated bridging helix harboring the communication module, which drives proper coordination of the catalytic residues. We then show the utility of CRISPR-integrated D43 in EL4 lymphocytes to regulate programmed cell death protein 1 (PD-1), a key receptor of immune checkpoints. Treatment of these cells with Tc showed a dose-dependent reduction in PD-1 by immunostaining and a decrease in messenger RNA levels by quantitative PCR as compared with wild type. PD-1 expression was recoverable upon removal of Tc. These results provide mechanistic insight into RNA devices with potential for cancer immunotherapy or other applications.