Mediator dynamics during heat shock in budding yeast.

Abstract

The Mediator complex is central to transcription by RNA polymerase II (Pol II) in eukaryotes. In budding yeast (Saccharomyces cerevisiae), Mediator is recruited by activators and associates with core promoter regions, where it facilitates preinitiation complex (PIC) assembly, only transiently before Pol II escape. Interruption of the transcription cycle by inactivation or depletion of Kin28 inhibits Pol II escape and stabilizes this association. However, Mediator occupancy and dynamics have not been examined on a genome-wide scale in yeast grown in nonstandard conditions. Here we investigate Mediator occupancy following heat shock or CdCl2 exposure, with and without depletion of Kin28. We find that Pol II occupancy shows similar dependence on Mediator under normal and heat shock conditions. However, although Mediator association increases at many genes upon Kin28 depletion under standard growth conditions, little or no increase is observed at most genes upon heat shock, indicating a more stable association of Mediator after heat shock. Unexpectedly, Mediator remains associated upstream of the core promoter at genes repressed by heat shock or CdCl2 exposure whether or not Kin28 is depleted, suggesting that Mediator is recruited by activators but is unable to engage PIC components at these repressed targets. This persistent association is strongest at promoters that bind the HMGB family member Hmo1, and is reduced but not eliminated in hmo1Δ yeast. Finally, we show a reduced dependence on PIC components for Mediator occupancy at promoters after heat shock, further supporting altered dynamics or stronger engagement with activators under these conditions.