ABORTIVE INITIATION AND FIRST BOND FORMATION AT AN ACTIVATED ADENOVIRUS E4 PROMOTER

Abortive initiation at the adenovirus E4 promoter was studied by follo wing the production of RNA formed from the initiating nucleotides UpA and CTP. Formation of a specific short RNA via a reaction with appropr iate alpha-amanitin sensitivity required promoter, activator, and ATP, In the absence of any of these, an alpha-amanitin-resistant reaction led to lower levels of a product of unknown origin. The alpha-amanitin -sensitive reaction required open promoter complexes, as assayed direc tly by permanganate probing. This reaction was not blocked by the inhi bition of polymerase C-terminal domain kinase activity or by the lack of DNA supercoiling. Thus, formation of the initial bond of the mRNA a ppears to require activator and ATP to open the DNA but not phosphoryl ation of the polymerase C-terminal domain, In addition, the abortive i nitiation reaction was strongly suppressed when all elongation substra tes were present, suggesting that cycling to produce high amounts of a bortive product is strongly disfavored during productive initiation at this promoter.