MicroRNA-sensing plasmid system for dynamic control of functional ion channel expression.

Gene therapy offers the potential for long-term treatment of a range of chronic diseases. However, permanent gene therapy expression may not be desirable. Efforts have been made to create systems that can be switched on/off by stimuli including light, designer drugs, or cellular contexts including increased electrical activity. Here, we designed a plasmid system in which ion channel expression and function are regulated by microRNA (miR)-an endogenous class of short noncoding RNAs which negatively regulate gene expression. We modified an existing voltage-gated potassium channel gene therapy with a binding cassette for miR-193a-3p, and transfected this "miR-193-OFF" system in neuro2A cells. Co-transfection with an inhibitor or mimic of miR-193a-3p, respectively, enhanced or repressed expression of our transgene, assessed using a GFP marker. Using whole-cell voltage clamp, we observed tuneable voltage-gated potassium currents in cells co-transfected with a miR-193a-3p inhibitor/mimic, compared with a non-targeting control oligonucleotide. Together, this demonstrates the concept of a miR-mediated molecular switch which can bias therapeutic ion channel expression based on a specific miR signal. As miRs are a ubiquitous molecular mechanism, our approach could be applied to a wide range of cellular and disease contexts, potentially expanding gene therapy to new patient populations.