Title: A reconstitution method towards characterizing the Mediator-ERa interaction for potential therapeutics
Speaker: Asst. Prof. Murat Alper Cevher (Bilkent University)
Time/location: March 27th, Wednesday at 15:40
Host: Bahar Değirmenci Uzun
Transcriptional activation entails a series of factors, including the class designated as coactivators. Of these, the most critical 30-subunit Mediator complex appears to be involved in the regulation of all RNA Polymerase II (Pol II) target genes and has emerged as the key integrative componant for transcription. Mediator action entails interactions of distinct subunits (for example, MED1 has been identified as one of the critical targets of nuclear receptors (NR), including ER whereas the MED17 is a target of p53 and MED15 is a target of multiple activators, including SREBP1, that are involved in fatty acid metabolism). Due to the massive size of the Mediator and the resulting technical limitations in dissecting its mechanisms of action, detailed activation pathways entailing this coactivator have not been elaborated yet. However, because the subunits are organized into discrete head, middle, tail, and kinase modules, a systematic structure-function approach is nonetheless feasible. We are currently reconstituting the ERa responsive form of the human Mediator complex in insect cells using the state-of-the-art Multibac baculovirus expression system. As a result of this reconstitution, we are now in a better shape in understanding how the Mediator interacts with ERa and recruit the general transcription machinery including the Pol II to the promoters. Our long term goal is to elucidate the interaction surface of ER-Mediator through biochemical as well as structural characterization, and to fully understand the recruitment mechanisms of the entire transcription machinery to ERa target genes with the Mediator. As different activators bind to different subunits/domains on the Mediator complex, by identifying ER-Mediator interface, we will be able to design specific inhibitors that will block ER-Mediator pathway and reduce side-affects and resistance issues that are observed with other therapeutics.