Chemical Engineering Seminar

Integrating synthetic circuitry into larger transcriptional networks to mediate predictable cellular behaviors remains a challenge within synthetic biology. In particular, the stochastic nature of transcription makes coordinating expression across multiple genetic elements difficult. Further, delivery of large genetic cargoes limits the efficiency of cellular engineering. Thus, our work is focused on the design of highly-compact genetic tools with a minimal genomic footprint. Co-localization of multiple transcriptional units provides a simple method of compact design. However, co-localization introduces the potential for physical coupling between transcriptional units. To address this challenge, we recently developed a theoretical framework for exploring how DNA supercoils—dynamic structures induced during transcription—influence transcription and gene expression in synthetic and native gene systems. Using this model, we find that DNA supercoiling strongly influences the profile of gene expression and that influence is defined by syntax—the relative orientation and position of genetic elements—and the enclosing boundary conditions. In exploring both synthetic and native gene regulatory networks, we find that supercoiling-mediated feedback changes the behaviors accessible to control and supports (or inhibits) the function of transcriptional networks. Importantly, we have recently confirmed several predictions from this model experimentally and used this model to design circuits with massively improved performance in primary cells. Our results suggest that supercoiling couples behavior between neighboring genes, representing a novel regulatory mechanism. Additionally, our predictions suggest why some circuit designs fail and provide a path to improving transgenic designs. Harnessing the insights from our model will enable enhanced transcriptional control, providing a robust method to tune expression levels, dynamics, and noise needed for the construction of transgenic systems for diverse cell engineering applications including cellular reprogramming.