Posttranslational Remote Control Mediated by Physical Inducers for Rapid Protein Release in Engineered Mammalian...

Physical cues such as light, heat, or an electrical field can be utilized for traceless, on-demand activation of the expression of a desired therapeutic gene in appropriately engineered cells with excellent spatiotemporal resolution, good inducibility, and simple reversibility. A similar approach can be applied to build a depolarization-based protein secretion system that enables rapid release of a therapeutic protein pre-stored in intracellular vesicles in mammalian cells. Here, we present a protocol to create designer β-cells that exhibit light-controllable rapid release (within 15 min) of a pre-synthesized proinsulin-nanoluciferase construct from vesicular stores. The construct is cleaved extracellularly to afford secreted insulin as a therapeutic protein and nanoluciferase as a reporter molecule. Such posttranslational remote control offers a much faster response than expression-based systems.