Mediator is a key regulator of eukaryotic transcription1, connecting activators and repressors bound to regulatory DNA elements with RNA polymerase II1,2,3,4 (Pol II). In the yeast Saccharomyces cerevisiae, Mediator comprises 25 subunits with a total mass of more than one megadalton (refs 5, 6) and is organized into three modules, called head, middle/arm and tail7,8,9. Our understanding of Mediator assembly and its role in regulating transcription has been impeded so far by limited structural information. Here we report the crystal structure of the essential Mediator head module (seven subunits, with a mass of 223 kilodaltons) at a resolution of 4.3 ångströms. Our structure reveals three distinct domains, with the integrity of the complex centred on a bundle of ten helices from five different head subunits. An intricate pattern of interactions within this helical bundle ensures the stable assembly of the head subunits and provides the binding sites for general transcription factors and Pol II. Our structural and functional data suggest that the head module juxtaposes transcription factor IIH and the carboxy-terminal domain of the largest subunit of Pol II, thereby facilitating phosphorylation of the carboxy-terminal domain of Pol II. Our results reveal architectural principles underlying the role of Mediator in the regulation of gene expression.
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We thank L. Messerle for providing the Ta6Br14 metal cluster, T. Hurley for reading the manuscript, M. Georgiadis for discussions, C. Kaplan for his advice on yeast genetics, T. Earnest for giving us beam time and L. Fabrizio for assisting with the N-terminal sequence analysis. We thank the CCP4 summer school, funded by the NCI (Y1-CO-1020), and NIGMS (Y1-GM-1104) for their assistance with the twinning data analysis. Y.T. thanks the instructors on ‘The X-Ray Methods Course’ at Cold Spring Harbor Laboratory. This work was supported by US National Science Foundation grant MCB 0843026 (Y.T.); the American Heart Association 0735395N (Y.T.); a Human Frontier Science Program long-term fellowship (T.I.); NIH grants R01 GM67167 (F.J.A.) and GM36659 (R.D.K.); NCI Cancer Center Support Grant P30 CA08748 (to the MSKCC Microchemistry and Proteomics Core Laboratory); and the European Commission Framework Program 7 projects INSTRUCT and P-CUBE (I.B.). X-ray data were collected at the GM/CA-CAT at the Advanced Photon Source, Argonne National Laboratory. GM/CA-CAT is funded by the NIH (Y1-CO-1020 and Y1-GM-1104) and the Advanced Photon Source is supported by the DOE (DE-AC02-06CH11357). Portions of this research were carried out at the Stanford Synchrotron Radiation Lightsource, supported by the DOE and the NIH, and at the Advanced Light Source, supported by the DOE (DE-AC02-05CH11231).
The authors declare no competing financial interests.
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Imasaki, T., Calero, G., Cai, G. et al. Architecture of the Mediator head module. Nature 475, 240–243 (2011). https://doi.org/10.1038/nature10162
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