(466e) Programmable Protein Circuits in Living Cells

Synthetic protein-­level circuits could enable engineering of powerful new cellular behaviors, complementary to the current circuits that predominantly use gene regulation and especially transcriptional regulation. Rational protein circuit design would be facilitated by a composable protein-protein regulation system, in which individual protein components can regulate one another in a "plug­-and-­play" fashion to create a variety of different circuit architectures. Here, we show that engineered viral proteases can function as composable protein components, which can together implement a broad variety of circuit-­level functions in mammalian cells (Gao, Chong, et al., 2018). In this system, termed CHOMP (Circuits of Hacked Orthogonal Modular Proteases) input proteases dock with and cleave target proteases to inhibit their function. These components can be connected to generate regulatory cascades, binary logic gates, analog filters, and dynamic signal­ processing functions. To demonstrate the potential biomedical utility of this system, we rationally designed a circuit that induces cell death in response to upstream activators of the Ras oncogene. Because CHOMP circuits can perform complex functions yet be encoded as single transcripts and delivered without genomic integration, they offer a scalable platform to facilitate protein circuit engineering for biotechnological applications. We will also discuss our recent efforts to expand the capabilities of our protein circuit platform, such as building more sensors for endogenous pathways, mining more orthogonal proteases as building blocks, and exploring a novel viral vector for circuit delivery.