Promoter Activation via a Cyclic AMP Response Element in Vitro

Transcription activation via activating transcription factor cyclic AMP response element binding (ATF/CREB) sites in vitro was explored using transcription and permanganate assay for open complex formation. These sites were used to drive transcription from an adenovirus major late core sequence. Under conditions where activation is strong, 20-50-fold, ATF/CREB is required for preinitiation complexes to reach the open complex stage. Complete opening requires activator, ATP, and initiating nucleotides. In exploration of postinitiation steps, no stimulation of promoter clearance was observed but a modest stimulation of the rate of continuous transcription occurred. High amounts of DNA template, commonly used in in vitro studies, allows some templates to open without activator, but leaves the nucleotide requirements intact. This leads to a drastic lowering of the dependence on ATF/CREB. Taken together, the data indicate that ATF/CREB activates this system primarily by stimulating the formation of functional preinitiation complexes.