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Synthetic translational regulation by an L7Ae–kink-turn RNP switch

Nature Chemical Biology volume 6, pages 71–78 (2010)Cite this article

Abstract

The regulation of cell signaling pathways and the reconstruction of genetic circuits are important aspects of bioengineering research. Both of these goals require molecular devices to transmit information from an input biomacromolecule to the desired outputs. Here, we show that an RNA-protein (RNP)-containing L7Ae–kink-turn interaction can be used to construct translational regulators under control of an input protein that regulates the expression of desired output proteins. We built a system in which L7Ae, an archaeal ribosomal protein, regulates the translation of a designed mRNA in vitro and in human cells. The translational regulator composed of the RNP might provide new therapeutic strategies based on the detection, repair or rewiring of intrinsic cellular defects, and it may also serve as an invaluable tool for the dissection of the behavior of complex, higher-order circuits in the cell.

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Figure 1: Schematic representation of the translational regulation research using the L7Ae–K-turn complex.
Figure 2: In vitro translational repression using the L7Ae–K-turn interaction.
Figure 3: In vitro translational activation using the L7Ae–K-turn interaction.
Figure 4: Translational repression in human cells.
Figure 5: Translational repression using an input protein encoded in the genome.

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Acknowledgements

The authors thank Y. Shimizu (University of Tokyo) for providing us with the PURE system solutions, A. Huttenhofer (Innsbruck Medical University, Biocenter) and T.S. Rozhdestvensky (University of Muenster) for providing us with the L7Ae plasmids, and S. Kashida (Kyoto University) for providing us with the p481 and pSk-7 plasmids. The authors also thank A. Kitamura (JST) for RNP structural modeling. This work was supported by a Grant-in-Aid for Young Scientists (A) (H.S.) and the International Cooperative Research Project, Japan Science and Technology Agency (H.S. and T.I.).

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Authors and Affiliations

  1. Laboratory of Gene Biodynamics, Graduate School of Biostudies, Kyoto University, Oiwake-cho, Kitashirakawa, Sakyo-ku, Kyoto, Japan

    Hirohide Saito, Tetsuhiro Kobayashi, Tomoaki Hara & Tan Inoue

  2. International Cooperative Research Project (ICORP), Japan Science and Technology Agency (JST), Sanban-cho, Chiyoda-ku, Tokyo, Japan

    Hirohide Saito, Yoshihiko Fujita, Karin Hayashi, Rie Furushima & Tan Inoue

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  1. Hirohide Saito
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  2. Tetsuhiro Kobayashi
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Contributions

H.S., T.K. and T.I. designed the experiments and analyzed the results. H.S., T.K., T.H. and R.F. performed the in vitro translation assays and the RNP interaction assays. Y.F. and K.H. performed the in vivo translation assays. H.S. and T.I. wrote the manuscript.

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Correspondence to Hirohide Saito or Tan Inoue.

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Saito, H., Kobayashi, T., Hara, T. et al. Synthetic translational regulation by an L7Ae–kink-turn RNP switch. Nat Chem Biol 6, 71–78 (2010). https://doi.org/10.1038/nchembio.273

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