Artificial receptors for mammalian synthetic biology

Synthetic biology has made great progress in engineering various components of cell signaling. These approaches are particularly relevant for generating therapeutic cells. The focus of this thesis concerns the development of tools for sensing biotechnologically or therapeutically important stimuli, such as disease markers, and for reprogramming cell behavior in response. The generalized extracellular molecule sensor (GEMS) design is based on the use of antibody fragments as ligand binding domains. The simultaneous binding of two such domains to a ligand causes the extracellular receptor domains to dimerize. This dimerization leads to a reorientation of the intracellular receptor domains, activating cellular signaling pathways. GEMS enabled the generation of cells that sense the cancer biomarker prostate-specific antigen (PSA) and activate transgene expression in response. The GEMS design was further used to construct receptors that sense heart damage. Cardiac troponin I is released by damaged cardiomyocytes, for example after acute myocardial infarction. Receptors that activate signaling pathways associated with tissue regeneration upon binding cardiac troponin I, may be a promising strategy for the development of cell therapy applications for heart disease. For some applications of artificial receptors, activating endogenous signaling pathways might cause unintended side effects. Therefore, we developed a synthetic signaling pathway based on phosphorylated histidine and aspartate residues in mammalian cells. This system is unlikely to interfere with endogenous signal processing, as the regulation of protein function by phosphorylation in mammalian cells is almost exclusively regulated by phosphorylated tyrosine, threonine and serine. All projects in this thesis provide insights into receptor development for different applications, and also enrich the toolbox of synthetic biology for cell signaling