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Mapping the fine structure of a eukaryotic promoter input-output function

Nature Genetics volume 45, pages 1207–1215 (2013)Cite this article

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Abstract

The precise tuning of gene expression levels is essential for the optimal performance of transcriptional regulatory networks. We created 209 variants of the Saccharomyces cerevisiae PHO5 promoter to quantify how different binding sites for the transcription factor Pho4 affect its output. We found that transcription-factor binding affinities determined in vitro could quantitatively predict the output of a complex yeast promoter. Promoter output was precisely tunable by subtle changes in binding-site affinity of less than 3 kcal mol−1, which are accessible by modifying 1–2 bases. Our results provide insights into how transcription-factor binding sites regulate gene expression, their possible evolution and how they can be used to precisely tune gene expression. More generally, we show that in vitro binding-energy landscapes of transcription factors can precisely predict the output of a native yeast promoter, indicating that quantitative models of transcriptional regulatory networks are feasible.

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Figure 1: Design and assembly of the promoter library.
Figure 2: On-chip induction, imaging and analysis.
Figure 3: Induction traces for each promoter in the library under Pi starvation.
Figure 4: Single Pho4 site variants.
Figure 5: Effects of split sites, dual site modifications and ablations.
Figure 6: Correlation between in vitro binding-site affinity and in vivo promoter output.
Figure 7: Phosphate-dependent activation of the PHO5 promoter.
Figure 8: Promoter dynamic range and induction threshold.

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Acknowledgements

We thank J.L. Garcia-Cordero for help with modeling phosphate uptake and the Shore lab at the University of Geneva for their assistance with the library construction. This work was supported by a SystemsX.ch grant DynamiX-RTD (2008/005) to S.J.M. and the École Polytechnique Fédérale de Lausanne.

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  1. Institute of Bioengineering, School of Engineering, Ecole Polytechnique Federale de Lausanne, Lausanne, Switzerland

    Arun S Rajkumar, Nicolas Dénervaud & Sebastian J Maerkl

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  1. Arun S Rajkumar
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Contributions

S.J.M. and A.S.R. designed the library constructs. A.S.R. synthesized the promoter libraries, performed nucleosome occupancy measurements and characterized the promoter library in bulk. N.D. carried out the on-chip experiments. N.D. and S.J.M. designed and implemented the transcription model. A.S.R. and S.J.M. analyzed the data. A.S.R. and S.J.M. wrote the manuscript.

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Correspondence to Sebastian J Maerkl.

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The authors declare no competing financial interests.

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Rajkumar, A., Dénervaud, N. & Maerkl, S. Mapping the fine structure of a eukaryotic promoter input-output function. Nat Genet 45, 1207–1215 (2013). https://doi.org/10.1038/ng.2729

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