Abstract
The Mediator complex of Saccharomyces cerevisiae is required for diverse aspects of transcription by RNA polymerase II (pol II). Mediator is composed of two functionally distinct subcomplexes, Rgr1 and Srb4. To identify the structures and functions of each subcomplex, we expressed recombinant proteins for each subunit and assayed their interactions with each other and with basal transcription proteins. The Rgr1 subcomplex is composed of the Gal11 module, which binds activators, and the Med9/10 module. The Med9/10 module is required for both transcriptional activation and repression, and these activities appear to be carried out by two submodules. Proteins in the Med9 submodule interact physically and genetically with Srb10/11, suggesting that the Med9 submodule mediates the repression of pol II. Purified recombinant Srb4 subcomplex stimulated basal transcription of pol II but had little effect on activated transcription and phosphorylation of the C-terminal domain of the Rpb1 subunit of pol II. Both subcomplexes of Mediator interacted with a distinct set of basal transcription factors and pol II. The modular organization of Mediator and the associated functions suggest that the Mediator complex may recruit and/or stabilize the preinitiation complex through several points of contact with transcriptional regulators and basal transcription factors.
Highlights
MRNA synthesis requires pol1 II and a set of general transcription factors (GTFs) including TFIIA, TFIIB, TFIID, TFIIF, and TFIIH
Taken together with the previous results, these findings suggested that the Rgr1 subcomplex contains Rgr1 plus the Gal11 module (Gal11, Sin4, Hrs1, and Med2), Med1, Med4, Med7, Med9, Med10, and Srb7, whereas the Srb4 subcomplex contains the remaining Mediator subunits, Srb2, Srb4, Srb5, Srb6, Rox3, Med6, Med8, and Med11
The Mediator complex is required for diverse aspects of the transcription process, such as activation, repression, and stimulation of basal transcription
Summary
Tion and the phosphorylation of the C-terminal domain (CTD) of the largest subunit of pol II (Rpb1) [6] To accommodate these diverse activities, the Mediator complex is composed of more than 20 polypeptides, including Srb subunits (Srb2, -4, -5, -6, and -7), Med subunits (Med1, -2, -4, -6, -7, -8, -9, -10, and -11), and several transcriptional coregulator subunits (Gal, Rgr, Sin, Hrs, and Rox3) (10 –17). In vitro binding assays have identified the Gal module as the activator binding target of the Mediator complex These results suggest that the Gal module is required for efficient recruitment of h-pol II to a promoter via activator-specific interaction during transcriptional activation. The biochemical function of the Srb subcomplex and the points of
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