Conservation of Promoter Melting Mechanisms in Divergent Regions of the Single-Subunit RNA Polymerases

The single-subunit RNA polymerases make up a widespread family of proteins found in phage, mitochondria, and chloroplasts. Unlike the phage RNAPs, the eukaryotic RNAPs require accessory factors to melt their promoters and diverge from the phage RNAPs in the regions where functions associated with promoter melting in the latter have been mapped, suggesting that promoter melting mechanisms in the eukaryotic RNAPs diverge from those in the phage enzymes. However, here we show that an element in the yeast mitochondria! RNAP, identified by sequence alignment with the T7 phage RNAP, fulfills a role in promoter melting similar to that filled by the T7RNAP intercalating hairpin. The yeast mitochondria! RNAP intercalating hairpin appears to be as important in promoter melting as the mitochondrial transcription factor, MTF1, and both a structurally integral hairpin and MTF1 are required to achieve high levels of transcription on a duplex promoter. Deletions from the hairpin also relieve MTF1 inhibition of promoter escape on premelted promoters, likely because such deletions disrupt interactions with the upstream edge of the transcription bubble. These results are consistent with recent structural and functional studies of human mitochondrial RNAP and further reveal the surprising extent of mechanistic conservation between the eukaryotic and phage-encoded members of the single-subunit RNAP family.