Abstract
Eukaryotic transcriptional repressors function by recruiting large coregulatory complexes that target histone deacetylase enzymes to gene promoters and enhancers. Transcriptional repression complexes, assembled by the corepressor NCoR and its homolog SMRT, are crucial in many processes, including development and metabolic physiology. The core repression complex involves the recruitment of three proteins, HDAC3, GPS2 and TBL1, to a highly conserved repression domain within SMRT and NCoR. We have used structural and functional approaches to gain insight into the architecture and biological role of this complex. We report the crystal structure of the tetrameric oligomerization domain of TBL1, which interacts with both SMRT and GPS2, and the NMR structure of the interface complex between GPS2 and SMRT. These structures, together with computational docking, mutagenesis and functional assays, reveal the assembly mechanism and stoichiometry of the corepressor complex.
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Acknowledgements
We thank P. Moody, P. Elliot and the beamline staff at DIAMOND and the European Synchrotron Radiation Facility for help with data collection. This work was supported by the Wellcome Trust (085408) and the UK Medical Research Council.
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The contributions of J.O., L.F. and P.J.W. were crucial to the final manuscript and these authors should be considered co–first authors. J.O. (assisted by J.T.G.) performed most of the protein cloning, expression, purification and interaction mapping, although important preliminary experiments were performed by B.C.K. J.O. prepared the GPS2–SMRT complex for NMR structure determination, which was carried out by J.-C.Y. and D.N. Crystallizations were performed primarily by J.O., with some later trials by J.A.G. and L.F. Crystal structures were determined by L.F., J.O. and J.W.R.S. The interaction motifs in GPS2 and SMRT were identified by J.W.R.S. and tested in pull-down experiments by J.O. Fluorescence polarization, coimmunoprecipitation and cotransfection-purification assays were performed by P.J.W. The two-hybrid assays, gel filtrations and NMR comparisons of wild-type and mutant TBL1 were performed by P.J.W., Z.C. and B.T.G. In silico docking experiments were performed by T.K. The laboratories of L.N. and D.N. provided experimental expertise for transfection and NMR studies, respectively. J.W.R.S. planned and supervised the project and prepared the manuscript with assistance from the other authors.
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Oberoi, J., Fairall, L., Watson, P. et al. Structural basis for the assembly of the SMRT/NCoR core transcriptional repression machinery. Nat Struct Mol Biol 18, 177–184 (2011). https://doi.org/10.1038/nsmb.1983
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DOI: https://doi.org/10.1038/nsmb.1983
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