Genetic circuit designs to improve synthetic biological signaling in mammalian cells

Multicellular organisms perform sophisticated computations on a myriad of environmental cues and produce many discrete cell states in response. These phenotypes are often defined by their type and level of gene expression. To recapitulate natural systems and engineer synthetic versions, the Computation via Recombinase Assisted Transcriptional Effectors (CREATE) platform presented here couples programmable transcription factors and site-specific recombinases to bridges an existing gap between digital and analog signal transmission in mammalian cells. This work demonstrates differential regulation of multiple protein expression levels in the same cell and complex operations, such as multiplication of two 2-bit recombinase input signals and convolution of digital and analog outputs in response to multiple inputs in human embryonic HEK293FT cells. To improve and predict signal transduction through biological circuits such as CREATE, a characterization framework is established for the quantitative assessment of recombinase digitizer modules that can be used to predict amplification of weak or leaky signals. This work demonstrates a quantitative improvement in biological signal transduction and robust mixed signal computation using CREATE in mammalian cells, with applications in drug screening, developmental biology, and tissue engineering.2024-05-23T00:00:00