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The glucocorticoid receptor dimer interface allosterically transmits sequence-specific DNA signals

Nature Structural & Molecular Biology volume 20, pages 876–883 (2013)Cite this article

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Abstract

Glucocorticoid receptor (GR) binds to genomic response elements and regulates gene transcription with cell and gene specificity. Within a response element, the precise sequence to which the receptor binds has been implicated in directing its structure and activity. Here, we use NMR chemical-shift difference mapping to show that nonspecific interactions with bases at particular positions in the binding sequence, such as those of the 'spacer', affect the conformation of distinct regions of the rat GR DNA-binding domain. These regions include the DNA-binding surface, the 'lever arm' and the dimerization interface, suggesting an allosteric pathway that signals between the DNA-binding sequence and the associated dimer partner. Disrupting this pathway by mutating the dimer interface alters sequence-specific conformations, DNA-binding kinetics and transcriptional activity. Our study demonstrates that GR dimer partners collaborate to read DNA shape and to direct sequence-specific gene activity.

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Figure 1: Nonspecific GBS bases modulate GR structure and activity.
Figure 2: GBS spacer affects the conformation of the D-loop.
Figure 3: Disruption of the dimerization interface affects lever arm and DNA-recognition helix conformation.
Figure 4: A477T impairs dimerization but not monomer DNA binding.
Figure 5: A477T disrupts cooperativity and GBS-specific dissociation.
Figure 6: Sequence-specific lever-arm conformation is dependent on the intact dimerization interface.

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Acknowledgements

We thank M. Kelly (University of California (UC), San Francisco) and J. Pelton (UC Berkeley) for NMR support, J. Cheney and S. Floor for providing scripts for NMR analysis, and S. Cooper and S. Meijsing for developing GR ChIP-sequencing protocols. We thank R. Fletterick, A. Johnson, G. Narlikar and members of K.R.Y.'s lab for thoughtful discussions; and M. Knuesel and S. Meijsing for critical reading of the manuscript. This work was supported by US National Institutes of Health (NIH) grant CA020535 (K.R.Y.), the Biophysics Graduate Group training grant NIHT32GM008284 (K.M.K.), NIH grants GM08537 and 5K99CA149088 (M.A.P.), the Leukemia and Lymphoma Society Fellowship (M.A.P.), the Larry S. Hillblom Foundation Fellowship (L.C.W.), the Genentech and Sandler Foundation Graduate Fellowship (L.C.W.) and the Cancer Research Coordinating Committee Fellowship (L.C.W.).

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  1. Miles A Pufall

    Present address: Present address: Department of Biochemistry, University of Iowa, Iowa City, Iowa, USA.,

Authors and Affiliations

  1. Tetrad Graduate Program, University of California, San Francisco, San Francisco, California, USA

    Lisa C Watson & Benjamin J Schiller

  2. Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, California, USA

    Lisa C Watson, Benjamin J Schiller, Miles A Pufall & Keith R Yamamoto

  3. Biophysics Graduate Group, University of California, San Francisco, San Francisco, California, USA

    Kristopher M Kuchenbecker

  4. Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, California, USA

    Kristopher M Kuchenbecker

  5. Department of Pharmaceutical Chemistry, University of California, San Francisco, San Francisco, California, USA

    John D Gross

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Contributions

L.C.W. designed, performed and analyzed experiments. J.D.G. and L.C.W. designed and performed experiments for NMR sequential assignment. K.M.K. and L.C.W. designed, performed and analyzed SPR and FRET experiments. B.J.S. and L.C.W. designed and performed ChIP-sequencing experiments, and B.J.S. analyzed the data. M.A.P. and K.R.Y. contributed to the direction of project, and L.C.W. and K.R.Y. wrote the manuscript.

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Correspondence to Keith R Yamamoto.

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Watson, L., Kuchenbecker, K., Schiller, B. et al. The glucocorticoid receptor dimer interface allosterically transmits sequence-specific DNA signals. Nat Struct Mol Biol 20, 876–883 (2013). https://doi.org/10.1038/nsmb.2595

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