Multiple layers of cooperativity regulate enhanceosome-responsive RNA polymerase II transcription complex assembly

Two coordinate forms of transcriptional synergy mediate eukaryotic gene reg ulation: the greater-than-additive transcriptional response to multiple pro moter-bound activators, and the sigmoidal response to increasing activator concentration. The mechanism underlying the sigmoidal response has not been elucidated but is almost certainly founded on the cooperative binding of a ctivators and the general machinery to DNA. Here,ve explore that mechanism by using highly purified transcription factor preparations and a strong Eps tein-Barr virus promoter, BHLF-1, regulated by the virally encoded activato r ZEBRA. We demonstrate that two layers of cooperative binding govern trans cription complex assembly. First, the architectural proteins HMG-1 and -2 m ediate cooperative formation of an enhanceosome containing ZEBRA and cellul ar Spl. This enhanceosome then recruits transcription factor IIA (TFIIA) an d TFIID to the promoter to form the DA complex, The DA complex, however, st imulates assembly of the enhanceosome itself such that the entire reaction can occur in a highly concerted manner. The data reveal the importance of r eciprocal cooperative interactions among activators and the general machine ry in eukaryotic gene regulation.