Abstract: The Mediator complex associates with eukaryotic RNA polymerase (Pol) II and is recruited to transcriptional enhancers by activator proteins. It is believed that Mediator is a general component of the Pol II machinery that is crucial to connect enhancer-bound activators to basic transcription factors. However, we show that Mediator does not detectably associate with many highly active Pol II promoters in yeast cells. Furthermore, in response to stress conditions, Mediator association is not directly related to Pol II association and in some cases is not detectable at highly activated promoters. Thus, Mediator is recruited to enhancers in an activator-specific manner, and it does not seem to be a stoichiometric component of the basic Pol II machinery in vivo. Mediator is recruited by many activators involved in stress responses, but not by the major activators that function under optimal conditions.