Rewiring native post-transcriptional global regulators to achieve designer, multi-layered genetic circuits

Trevor R. Simmons, Gina Partipilo, Ryan Buchser, Anna C. Stankes, Rashmi Srivastava, Darian Chiu, Benjamin K. Keitz and Lydia M. Contreras ()
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Trevor R. Simmons: The University of Texas at Austin
Gina Partipilo: The University of Texas at Austin
Ryan Buchser: The University of Texas at Austin
Anna C. Stankes: The University of Texas at Austin
Rashmi Srivastava: Baylor University
Darian Chiu: The University of Texas at Austin
Benjamin K. Keitz: The University of Texas at Austin
Lydia M. Contreras: The University of Texas at Austin

Nature Communications, 2024, vol. 15, issue 1, 1-13

Abstract: Abstract As synthetic biology expands, creating “drag-and-drop” regulatory tools that can achieve diverse regulatory outcomes are paramount. Herein, we develop a approach for engineering complex post-transcriptional control by rewiring the Carbon Storage Regulatory (Csr) Network of Escherichia coli. We co-opt native interactions of the Csr Network to establish post-transcriptional logic gates and achieve complex bacterial regulation. First, we rationally engineer RNA-protein interactions to create a genetic toolbox of 12 BUFFER Gates that achieves a 15-fold range of expression. Subsequently, we develop a Csr-regulated NOT Gate by integrating a cognate 5’ UTR that is natively Csr-activated into our platform. We then deploy the BUFFER and NOT gates to build a bi-directional regulator, two input Boolean Logic gates OR, NOR, AND and NAND and a pulse-generating circuit. Last, we port our Csr-regulated BUFFER Gate into three industrially relevant bacteria simply by leveraging the conserved Csr Network in each species.

Date: 2024
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DOI: 10.1038/s41467-024-52976-1

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