The S. cerevisiae RNA polymerase III (pol III) transcription factor TFIIIB binds to DNA upstream of the transcription start site of the SUP4 tRNA(Tyr) gene in a TFIIIC-dependent reaction and to the major 5S rRNA gene in a reaction requiring TFIIIC and TFIIIA. It is shown here that TFIIIB alone correctly positions pol III for repeated cycles of transcription on both genes, with the same efficiency as fully assembled transcription complexes. Thus, TFIIIB is the sole transcription initiation factor of S. cerevisiae pol III; TFIIIC and TFIIIA are assembly factors for TFIIIB. The TFIIIB-dependent binding of pol III to the SUP4 tRNA and 5S rRNA genes has been analyzed in binary (protein and DNA only) and precisely arrested ternary (protein, DNA, and RNA) transcription complexes. Pol III unwinds at least 14 bp of DNA at the SUP4 transcription start in a temperature-dependent process. The unwound DNA segment moves downstream with nascent RNA as a transcription bubble of approximately the same size.
Base Sequence DNA, Fungal