A minimal RNA polymerase III transcription system.

Transcription factor (TF) IIIB recruits RNA polymerase (pol) III for specific initiation of transcription. All three subunits of TFIIIB, TBP, Brf (the TFIIB-related subunit) and B", are required for transcription of supercoiled and linear duplex DNA, but we show here that B" is non-essential on a promoter that has been partly pre-opened by unpairing a short segment of the transcription bubble. These findings expose a striking similarity between transcriptional initiation by pol II, pol III and bacterial RNA polymerases: a preformed single-stranded DNA bubble upstream of the transcriptional start removes the dependence of pol II on TFIIE, TFIIH and ATP hydrolysis, and the dependence of pol III on B"; the favored placement of the transcription bubble for B"-independent transcription by pol III overlaps a DNA segment that interacts sequence specifically as single-stranded DNA with the sigma(70 )initiation subunit of Escherichia coli RNA polymerase holoenzyme.