The behavior of gene modules in complex synthetic circuits is often unpredictable1,2,3,4. After joining modules to create a circuit, downstream elements (such as binding sites for a regulatory protein) apply a load to upstream modules that can negatively affect circuit function1,5. Here we devised a genetic device named a load driver that mitigates the impact of load on circuit function, and we demonstrate its behavior in Saccharomyces cerevisiae. The load driver implements the design principle of timescale separation: inclusion of the load driver's fast phosphotransfer processes restores the capability of a slower transcriptional circuit to respond to time-varying input signals even in the presence of substantial load. Without the load driver, we observed circuit behavior that suffered from a 76% delay in response time and a 25% decrease in system bandwidth due to load. With the addition of a load driver, circuit performance was almost completely restored. Load drivers will serve as fundamental building blocks in the creation of complex, higher-level genetic circuits.
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We thank members of the labs of R.W. and D.D.V. for discussions, M.-T. Chen (Department of Electrical Engineering, Princeton University) for plasmids containing both STAT5-HKRR and JAK2, and the Synthetic Biology Center at Massachusetts Institute of Technology's cytometry facility. D.M. was supported by the Eni-MIT Energy Research Fellowship, and both D.M. and P.M.R. were supported by the National Science Foundation (NSF) Graduate Research Fellowship Plan under grant DGE-1122374. This research was supported by the NSF (CCF-1058127), NSF SynBERC (SA5284-11210), USAFOSR (FA9550-12-1-0129), USARO ICB (W911NF-09-D-0001) and the US National Institutes of Health (P50 GM098792).
The authors declare no competing financial interests.
Supplementary Figures 1–24, Supplementary Tables 1–7 and Supplementary Note 1 (PDF 3671 kb)
Matlab files containing mathematical model implementation and code to plot all simulation and experimental results (Figs. 2 and 3, Supplementary Figs. 4-5, 7-18, and 23-24). (ZIP 19441 kb)
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Mishra, D., Rivera, P., Lin, A. et al. A load driver device for engineering modularity in biological networks. Nat Biotechnol 32, 1268–1275 (2014). https://doi.org/10.1038/nbt.3044
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