Structure of the Mediator head module

Abstract

Gene transcription by RNA polymerase (Pol) II requires the coactivator complex Mediator. Mediator connects transcriptional regulators and Pol II, and is linked to human disease1–4. Mediator from the yeast Saccharomyces cerevisiae has a molecular mass of 1.4 megadaltons and comprises 25 subunits that form the head, middle, tail and kinase modules5–7. The head module constitutes one-half of the essential Mediator core8, and comprises the conserved9 subunits Med6, Med8, Med11, Med17, Med18, Med20 and Med22. Recent X-ray analysis of the S. cerevisiae head module at 4.3A˚ resolution led to a partial architectural model with three submodules called neck, fixed jaw and moveable jaw10. Here we determine de novo the crystal structure of the head module from the fission yeast Schizosaccharomyces pombe at 3.4A˚ resolution. Structure solution was enabled by new structures of Med6 and the fixed jaw, and previous structures of the moveable jaw11 and part of the neck12, and required deletion of Med20. The S. pombe head module resembles the head of a crocodile with eight distinct elements, of which at least four are mobile. The fixed jaw comprises tooth and nose domains, whereas the neck submodule contains a helical spine and one limb, with shoulder, arm and finger elements. The arm and the essential shoulder contact other parts of Mediator. The jaws and a central joint are implicated in interactions with Pol II and its carboxy-terminal domain, and the joint is required for transcription in vitro. The S. pombe head module structure leads to a revised model of the S. cerevisiae module, reveals a high conservation and flexibility, explains knownmutations, and provides the basis for unravelling a central mechanism of gene regulation

Mediator links transcription and DNA repair by facilitating Rad2/XPG recruitment

“…We suggest that the contact between Med17 and Rad2 is direct, since our two-hybrid screening permitted us to reveal only specific interactions between different subunits within the Mediator complex (Guglielmi et al 2004) that were further confirmed as direct by structural analysis (for example, Lariviere et al 2012;Robinson et al 2012). Moreover, we demonstrated that several mutations in the Med17 Mediator subunit resulted in a decreased interaction between Mediator and Rad2, an observation in favor of a direct Mediator-Rad2 contact.…”

“…Protein-protein interactions within the Mediator complex have been published (Guglielmi et al 2004). Many interactions that we uncovered in the screen were confirmed crystallographically and served as a basis for other Mediator studies (for example, Lariviere et al 2012;Robinson et al 2012). In addition, our two-hybrid screening revealed a large number of interactions between S. cerevisiae Mediator and other nuclear proteins (B Guglielmi, NL van Berkum, C Boschiero, FCP Holstege, and M Werner, unpubl.).…”

“…The complete crystallographic structure of the Mediator complex has not been determined. However, recent structural data have been obtained for the Mediator head module (Lariviere et al 2012;Robinson et al 2012). Mediator subunits are engaged in numerous contacts within the complex (Guglielmi et al 2004) and probably with partners belonging to other components of the transcription machinery.…”

Mediator Recruitment to Heat Shock Genes Requires Dual Hsf1 Activation Domains and Mediator Tail Subunits Med15 and Med16

“…For example, the Head module plays an important role in the assembly and/or stabilization of the transcription initiation complex. Head subunits assemble into a jawlike structure that interacts with the CTD of the largest Pol II subunit, Rpb1, as well as with Rpb3 (7,8). The flexibility and extended shape of the Head permits interactions with other components of the general transcriptional machinery, including TBP, TFIIB, and TFIIH (8 -13).…”

Evidence for Multiple Mediator Complexes in Yeast Independently Recruited by Activated Heat Shock Factor

“…1A) contributes to its multiple layers of function. For example, the head module is assembled into a jawlike structure comprised of fixed (Med17-Med11-Med22) and movable jaws (the latter consisting of the C terminus of Med8 joined to the Med18-Med20 heterodimer) (13). The flexibility and extended shape of the head permit interactions with Pol II at three putative interaction surfaces-on Rpb1 (within its C-terminal domain [CTD]), Rpb3, and the Rpb4/Rpb7 subcomplex-as well as with other components of the transcription initiation complex, including TBP, TFIIB and TFIIH (13)(14)(15)(16)(17)(18).…”