A 3-STEP PATHWAY OF TRANSCRIPTION INITIATION LEADING TO PROMOTER CLEARANCE AT AN ACTIVATED RNA-POLYMERASE-II PROMOTER

The progress of transcription bubbles during initiation in vitro was f ollowed in order to learn how RNA polymerase II begins transcription a t the activated adenovirus E4 promoter, The issues addressed include t he multiple roles of ATP, the potential effect of polymerase C-termina l domain phosphorylation, and the ability of polymerase to clear the p romoter for reinitiation. The results lead to a three-step model for t he transition from closed complex to elongation complex, two steps of which use ATP independently, In the first step, studied previously, AT P is hydrolyzed to open the DNA strands over the start site. In a seco nd step, apparently independent of ATP, transcription bubbles move int o the initial transcribed region where RNA synthesis can stall. In the third step, transcripts can be made as polymerase is released from th ese stalled positions,vith the assistance of an ATP-dependent process, likely phosphorylation of the polymerase C-terminal domain, After thi s third step, tile promoter becomes cleared, allowing for the reinitia tion of transcription.