THE REQUIREMENT FOR THE BASAL TRANSCRIPTION FACTOR IIE IS DETERMINED BY THE HELICAL STABILITY OF PROMOTER DNA

The role of the basal transcription factor TFIIE was investigated in RNA polymerase II transcription reactions reconstituted with purified proteins. Using negatively supercoiled templates, which circumvent the requirement for TFIIH, we observed that transcription from the adenovirus major-late (ML) core promoter is more dependent on TFIIE than transcription from the adenovirus E4 (E4) or mouse mammary tumor virus (MMTV) promoters. For all three promoters, an increase in the ionic strength of the reaction mixtures led to an increased dependence on TFIIE. Analysis of hybrid ML/MMTV promoters showed that the region encompassing the start site, from -10 to +10, dictates this dependence. Transcription from a relaxed E4 template with a pre-melted -8 to +2 region was completely independent of both TFIIE and TFIIH. We propose that on negatively supercoiled templates TFIIE can facilitate promoter melting.