A load driver device for engineering modularity in biological networks

Abstract

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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Figure 1: Block diagrams of unbuffered and buffered systems.
Figure 2: Attenuation of retroactivity by faster load driver dynamics.
Figure 3: System responses to step inputs and dosage response.
Figure 4: System responses to periodic inputs.
Figure 5

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Acknowledgements

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).

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D.M., D.D.V. and R.W. designed the experiments and analyzed the data. D.M. performed the experiments. P.M.R. constructed mathematical models and performed parameter estimation. A.L. cloned constructs. D.M., D.D.V. and R.W. wrote the paper.

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Correspondence to Domitilla Del Vecchio or Ron Weiss.

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The authors declare no competing financial interests.

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Supplementary Text and Figures

Supplementary Figures 1–24, Supplementary Tables 1–7 and Supplementary Note 1 (PDF 3671 kb)

Supplementary Code

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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