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Disorder and residual helicity alter p53-Mdm2 binding affinity and signaling in cells

Nature Chemical Biology volume 10, pages 1000–1002 (2014)Cite this article

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Abstract

Levels of residual structure in disordered interaction domains determine in vitro binding affinities, but whether they exert similar roles in cells is not known. Here, we show that increasing residual p53 helicity results in stronger Mdm2 binding, altered p53 dynamics, impaired target gene expression and failure to induce cell cycle arrest upon DNA damage. These results establish that residual structure is an important determinant of signaling fidelity in cells.

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Figure 1: Transient p53TAD helicity determines the binding affinity to Mdm2.
Figure 2: Altered cellular dynamics of helical p53TAD mutants.
Figure 3: Altered target gene expression and cell-cycle profiles of helical p53TAD mutants.

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Acknowledgements

We thank P. Schmieder and M. Beerbaum for excellent NMR infrastructure maintenance (FMP Berlin) and W. Chen for Illumina sequencing (Max Delbrueck Center). This research was supported by the Deutsche Forschungsgemeinschaft (Emmy Noether grant PS1794/1-1 to P.S.), the Association pour la Recherche contre le Cancer (postdoctoral fellowship to F.-X.T.), the European Union FP7 (Marie Curie CIG to A.L.), the American Cancer Society (RSG-07-289-01-GMC to G.W.D.) and the National Science Foundation (MCB-0939014 to G.W.D.). Protein NMR resources at the University of South Florida were provided by the Center for Drug Discovery and Innovation.

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Author notes

  1. Wade Borcherds, François-Xavier Theillet and Andrea Katzer: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Cell Biology, Microbiology and Molecular Biology, University of South Florida, Tampa, Florida, USA

    Wade Borcherds, Katie M Mishall, Anne T Powell, Hongwei Wu & Gary W Daughdrill

  2. Center for Drug Discovery and Innovation, University of South Florida, Tampa, Florida, USA

    Wade Borcherds, Katie M Mishall, Anne T Powell, Hongwei Wu & Gary W Daughdrill

  3. Department of NMR-supported Structural Biology, In-cell NMR Laboratory, Leibniz Institute of Molecular Pharmacology (FMP Berlin), Berlin, Germany

    François-Xavier Theillet & Philipp Selenko

  4. Berlin Institute for Medical Systems Biology, Max Delbrueck Center for Molecular Medicine, Berlin, Germany

    Andrea Katzer, Ana Finzel & Alexander Loewer

  5. Drug Discovery Department, Moffitt Cancer Center, University of South Florida, Tampa, Florida, USA

    Wanda Manieri

  6. Max Planck Institute for Biology of Ageing, Cologne, Germany

    Christoph Dieterich

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  1. Wade Borcherds
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Contributions

W.B., F.-X.T. and H.W. collected and analyzed NMR data. W.B., K.M.M. and A.T.P. prepared p53TAD and Mdm2 protein samples, F.-X.T., K.M.M. and A.T.P. collected and analyzed ITC data. W.M. provided the protocol for Mdm2 expression and purification. F.-X.T. and A.K. generated and analyzed mutant p53 cell lines, A.F. performed RNA-seq experiments, and C.D. provided bioinformatics analysis. F.-X.T. and W.B. assisted in the writing of the manuscript. P.S., A.L. and G.W.D. conceived the study, supervised the research, analyzed the data, prepared the figures and wrote the manuscript.

Corresponding authors

Correspondence to Philipp Selenko, Alexander Loewer or Gary W Daughdrill.

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The authors declare no competing financial interests.

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Supplementary Results and Supplementary Figures 1–10. (PDF 19769 kb)

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Borcherds, W., Theillet, FX., Katzer, A. et al. Disorder and residual helicity alter p53-Mdm2 binding affinity and signaling in cells. Nat Chem Biol 10, 1000–1002 (2014). https://doi.org/10.1038/nchembio.1668

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