A Dynamic Model for the Function of Mediator Modules In Vivo

Mediator is a highly conserved transcriptional coactivator organized into four modules, namely Tail, Middle, Head and Kinase (CKM). Previous work suggests regulatory roles for Tail and CKM, but an integrated model for these activities is lacking. Here, we analyzed the genome-wide distribution of Mediator subunits in wild-type and mutant yeast cells in which RNA polymerase II promoter escape is blocked allowing detection of transient Mediator forms. We found that while all modules are recruited to upstream activated regions (UAS), assembly of Mediator within the pre-initiation complex is accompanied by the release of CKM. Surprisingly, our data show that CKM regulates Mediator-UAS interaction rather than Mediator-promoter association. In addition, while Tail is required for Mediator recruitment to UAS, Tail-less Mediator nevertheless interacts with core promoters. Collectively, our data suggest that the essential function of Mediator is mediated by Head and Middle at core promoters, while Tail and CKM play regulatory roles. Overall design: ChIP-exo of 2 subunits of mediator (MED15 and MED7) in Saccharomyces cerevisiae using MYC epitope tag in kin28as strains. Samples are done in replicates. 2 combined No-Tag samples were process and used to normalize the combined experiment sample (by subtraction). ChIP-exo of Sua7 (TFIIB) in Saccharomyces cerevisiae using TAP epitope tag. Samples are done in replicates. 4 combined No-Tag samples were process and used to normalize the combined experiment sample (by subtraction).