title: Suppression of Mediator is regulated by Cdk8-dependent Grr1 turnover of the Med3 coactivator.
abstract: Mediator, an evolutionary conserved large multisubunit protein complex with a central role in regulating RNA polymerase II-transcribed genes, serves as a molecular switchboard at the interface between DNA binding transcription factors and the general transcription machinery. Mediator subunits include the Cdk8 module, which has both positive and negative effects on activator-dependent transcription through the activity of the cyclin-dependent kinase Cdk8, and the tail module, which is required for positive and negative regulation of transcription, correct preinitiation complex formation in basal and activated transcription, and Mediator recruitment. Currently, the molecular mechanisms governing Mediator function remain largely undefined. Here we demonstrate an autoregulatory mechanism used by Mediator to repress transcription through the activity of distinct components of different modules. We show that the function of the tail module component Med3, which is required for transcription activation, is suppressed by the kinase activity of the Cdk8 module. Med3 interacts with, and is phosphorylated by, Cdk8; site-specific phosphorylation triggers interaction with and degradation by the Grr1 ubiquitin ligase, thereby preventing transcription activation. This active repression mechanism involving Grr1-dependent ubiquitination of Med3 offers a rationale for the substoichiometric levels of the tail module that are found in purified Mediator and the corresponding increase in tail components seen in cdk8 mutants.
authors.affiliation: College of Medicine, Swansea University, Swansea, Wales SA2 8PP, United Kingdom.
authors.forename: Deyarina
authors.initials: D
authors.lastname: Gonzalez
authors.forename: Nurul
authors.initials: N
authors.lastname: Hamidi
authors.forename: Ricardo
authors.initials: R
authors.lastname: Del Sol
authors.forename: Joris J
authors.initials: JJ
authors.lastname: Benschop
authors.forename: Thomas
authors.initials: T
authors.lastname: Nancy
authors.forename: Chao
authors.initials: C
authors.lastname: Li
authors.forename: Lewis
authors.initials: L
authors.lastname: Francis
authors.forename: Manuel
authors.initials: M
authors.lastname: Tzouros
authors.forename: Jeroen
authors.initials: J
authors.lastname: Krijgsveld
authors.forename: Frank C P
authors.initials: FC
authors.lastname: Holstege
authors.forename: R Steven
authors.initials: RS
authors.lastname: Conlan
mesh_terms: D047369:Chromatin Immunoprecipitation; D002853:Chromatography, Liquid; D056850:Cyclin-Dependent Kinase 8 / Q000378:metabolism*; D044783:F-Box Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000235:genetics*; D015151:Immunoblotting; D013058:Mass Spectrometry; D056892:Mediator Complex / Q000037:antagonists & inhibitors* / Q000235:genetics; D046228:Microarray Analysis; D010766:Phosphorylation; D060888:Real-Time Polymerase Chain Reaction; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D014158:Transcription, Genetic / Q000235:genetics / Q000502:physiology*; D020798:Two-Hybrid System Techniques; D044767:Ubiquitin-Protein Ligases / Q000378:metabolism*
chemical_list: D044783:F-Box Proteins; D056892:Mediator Complex; C110281:PGD1 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C071032:GRR1 protein, S cerevisiae; D044767:Ubiquitin-Protein Ligases; D056850:Cyclin-Dependent Kinase 8