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Engineering High-Precision, Dynamic Genetic Control Systems for Cellular Reprogramming

Katie Galloway (MIT)

Thursday, Feb 2, 2023

Integrating synthetic circuitry into larger transcriptional networks to mediate predictable cellular behaviors remains a challenge within synthetic biology. Rational de novo design of synthetic circuits for cell engineering remains challenging. In particular, the stochastic nature of transcription makes coordinating expression across multiple genetic elements challenging. To address this challenge, my lab 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. We find that gene syntax—the relative ordering and orientation of genes—defines the expression profiles, variance, burst dynamics, and intergene correlation of two-gene systems. By applying our model to both a synthetic toggle switch and the endogenous zebrafish segmentation network, we find that supercoiling can enhance or weaken conventional biochemical regulatory strategies such as mRNA- and protein-mediated feedback loops. In cell culture, we confirmed that two-gene circuits qualitatively match the syntax-specific profiles predicted by our model. Our model integrates supercoiling-mediated biophysical feedback with classic gene regulation motifs such as transcriptional repressors that are well-studied in native and synthetic contexts. Our model provides an extensible framework for modeling an arbitrary number of genes and regulatory architectures. 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 transgene systems including synthetic gene circuits in primary cells and diverse cell engineering applications including cellular reprogramming.

Speaker Bio

Katie Galloway, Ph.D. is the W. M. Keck Career Development Professor in Biomedical Engineering and Chemical Engineering at MIT. Her research focuses on elucidating the fundamental principles of integrating synthetic circuitry to drive cellular behaviors. Through a combination of systems and synthetic biology approaches, her lab develops integrated gene circuits and elucidates the systems-level principles that govern complex cellular behaviors. Her goal is to leverage quantitative tools to transform how we understand cellular transitions and engineer cellular therapies. Galloway earned a PhD and an MS in Chemical Engineering from the California Institute of Technology, and a BS in Chemical Engineering from University of California at Berkeley. She completed her postdoctoral work at University of Southern California Stem Cell. Her research has been featured in Science, Cell Stem Cell, Cell Systems, and Development. She has won multiple fellowships and awards including the NIH Maximizing Investigators' Research Award R35, the NIH F32, and Caltech’s Everhart Award. Together with her husband she’s raising four awesome rambunctious, tenacious kids who are 5, 8, 11, and 13.

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