(627g) A Platform Technology for Dynamic Control of Cell Behavior

Mammalian cells rely on complex and highly dynamic gene networks to maintain cellular homeostasis in response to environmental stimuli and intracellular signals. Efficient cellular reprogramming thus requires integration of exogenous components with endogenous cellular networks through feedback control systems. We explored the use of post-translational tools for superior feedback regulation of dynamic behaviors. Specifically, we demonstrated efficient detection and manipulation of the main cellular stress response system – the Unfolded Protein Response (UPR) – for the design of high producing cell lines for protein manufacturing and for the development of cell therapies for sustained protein production.

Our approach is based on the use of the NanoDeg (ACS Synth Biol. 2018 Feb 16;7(2):540-552) – a bifunctional system that mediates proteasomal degradation of a cellular target with high specificity and exquisite control over rate of decay. To achieve input-dependent post-translational control of the output through the NanoDeg, feedforward loop topologies were explored and compared to conventional strategies for circuit design. iterations of mathematical modeling and experimental tests allowed defining the ideal circuit architecture for integrating the NanoDeg within orthogonal cellular networks.

This work generated a platform technology for quantitative, multiplexed profiling of gene expression signatures of the UPR with high sensitivity and dynamic resolution of the stimulus causing proteotoxic stress. This study also provides the design rules of a novel cell engineering technology for building complex genetic networks that govern highly dynamic cellular behaviors.