Promoter Length Affects the Initiation of T7 RNA Polymerase In Vitro: New Insights into Promoter/Polymerase...

Polymerases are integral factors of gene expression and are essential for the maintenance and transmission of genetic information. RNA polymerases (RNAPs) differ from other polymerases in that they can bind promoter sequences and initiate transcription de novo and this promoter recognition requires the presence of specific DNA binding domains in the polymerase. Bacteriophage T7 RNA polymerase (T7RNAP) is the prototype for single subunit RNA polymerases which include bacteriophage and mitochondrial RNAPs, and the structure and mechanistic aspects of transcription by T7 RNAP are well characterized. Here, we describe experiments to determine whether the prototype T7 RNAP is able to recognize and initiate at truncated promoters similar to mitochondrial promoters. Using an in vitro oligonucleotide transcriptional system, we have assayed transcription initiation activity by T7 RNAP. These assays have not only defined the limits of conventional de novo initiation on truncated promoters, but have identified novel activities of initiation of RNA synthesis. We propose that these novel activities may be vestigial activities surviving from the transition of single subunit polymerase initiation using primers to de novo initiation using promoters.