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Rapid and orthogonal logic gating with a gibberellin-induced dimerization system

Abstract

Using a newly synthesized gibberellin analog containing an acetoxymethyl group (GA3-AM) and its binding proteins, we developed an efficient chemically inducible dimerization (CID) system that is completely orthogonal to existing rapamycin-mediated protein dimerization. Combining the two systems should allow applications that have been difficult or impossible with only one CID system. By using both chemical inputs (rapamycin and GA3-AM), we designed and synthesized Boolean logic gates in living mammalian cells. These gates produced output signals such as fluorescence and membrane ruffling on a timescale of seconds, substantially faster than earlier intracellular logic gates. The use of two orthogonal dimerization systems in the same cell also allows for finer modulation of protein perturbations than is possible with a single dimerizer.

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Figure 1: The gibberellin-inducible CID system functions on a timescale of seconds.
Figure 2: Gibberellin-based CID can induce localized protein activity and is orthogonal to rapamycin CID.
Figure 3: A fast-processing OR logic gate in living cells using two CID systems.
Figure 4: A fast-processing AND logic gate in living cells using two CID systems.

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Acknowledgements

This study was supported in part by the US National Institutes of Health (NIH; GM092930 and DK090868 to T.I. and NS072241 to M.J.W), the US National Science Foundation (IOS-0641548 and MCB-0923723 to T.S.) and the National Center for Research Resources of the NIH and NIH Roadmap for Medical Research (UL1 RR 025005 to C.M. and D.J.M). T.U. is a recipient of a fellowship from the Japanese Society for the Promotion of Science. M.C. is a recipient of the Provost's Undergraduate Research Award.

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Authors and Affiliations

Authors

Contributions

T.M., A.S., M.C. and T.I. generated DNA constructs and T.M., R.D., T.U., A.S. and T.I. carried out cell biology experiments. T.S. advised on design of the gibberellin system. T.M. conducted biochemical experiments under supervision of M.J.W. C.M. and D.J.M. synthesized GA3-AM and GA3-H. T.I. conceived ideas. R.D. and T.I. wrote the paper.

Corresponding author

Correspondence to Takanari Inoue.

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There is a pending patent associated with the gibberellin-induced dimerization system.

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Supplementary Movie 1

Supplementary Movie 1 (Miyamoto and DeRose et al.) (AVI 11905 kb)

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Miyamoto, T., DeRose, R., Suarez, A. et al. Rapid and orthogonal logic gating with a gibberellin-induced dimerization system. Nat Chem Biol 8, 465–470 (2012). https://doi.org/10.1038/nchembio.922

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