Post-translational control schemes in the regulation of synthetic cellular signaling via engineered notch receptors

The development of engineered and orthogonal biomolecular devices in mammalian cells has granted the ability to customize and tune the way in which cells sense and respond to stimuli. These synthetic signaling pathways have often taken inspiration from endogenous pathways, such as the use of transcription factors or induced nuclear translocation, and have been successful in generating customized cell therapies and advancing tissue engineering. However, one ubiquitous and dominant paradigm for endogenous cellular signaling that has not been fully realized in synthetic contexts is the use of the covalent attachment of small molecules to proteins. These post-translational modifications to proteins often function as an additional regulatory method – one that functions on top of traditional signaling – changing how cells sense and respond to stimuli dependent on their context. Here, we demonstrate the development of a synthetic and orthogonal post-translational modification for mammalian cellular signaling. Using the synthetic Notch receptor as a model pathway, we illustrate the ability for cells to sense and respond to inputs dependent on the modification state of receptors. We further describe the extension of these tools for generalized and inducible signaling of synthetic Notch receptors, as well as their use in generating modification-dependent translocation and transcription. By mimicking the methods in which endogenous pathways control their sensing abilities, engineered modification-dependent pathways are positioned to approach more complex signaling states than with traditional methods alone.2024-01-26T00:00:00