Protein circuits reprogram cells

Abstract

Two protein circuit systems, split-protease-cleavable orthogonal coiled-coil logic (SPOC logic) and circuits of hacked orthogonal modular proteases (CHOMP), have been developed to permit rapid and logic function-based control of mammalian cellular signaling.

Fig. 1 |. Engineering protein circuit networks for cellular programming.

a, The design of a protease-activatable reporter in SPOC logic. OFF state (left): a protease is split into two halves and fused to inducible dimers (cyan and yellow). Antiparallel coiled-coil pairs, typically consisting of an inhibitory (green) and a target coil (blue) are connected by a substrate linker (red) for the autoinhibition of reporter proteins before activation. ON state (right): the proteolytic activity can be inducible by a chemical inducer triggering the dimerization of pairs. Once the linker between the inhibitory and target coils is cleaved by the induced protease, a displacer coil (orange) can interact with the target coil to activate the reporter protein for detection. b, Boolean logic gates can be engineered to integrate multiple inputs and produce specific outputs. c, Protease-regulated proteases can be engineered to serve as the output of an upstream layer but the input of the next layer to propagate signals across multiple layers. d, The design of a protease-activatable reporter in CHOMP. A degron motif is fused to a citrine fluorescent protein reporter via a protease substrate linker. The induced protease cleavage at the targeting substrate site removes the degron to stabilize and activate the citrine reporter.