DNA Supercoiling Alleviates Cold-Sensitivity of Promoter Melting by Extremophilic Deinococcus-Thermus RNA Polymerases

Abstract

Melting of promoter DNA around the transcription start site (TSS) is a critical step of transcription required for initiation of RNA synthesis. In bacteria, promoter melting is mediated by the holoenzyme of RNA polymerase (RNAP) consisting of the catalytic core enzyme and the promoter recognition subunit, σ factor. Previously, we showed that RNAPs from thermophilic Thermus aquaticus and mesophilic Deinococcus radiodurans are unable to open promoters at ambient temperatures and require heating for DNA melting. These properties depend on their σ factors and are recapitulated in the hybrid holoenzymes including these σ factors and the core enzyme of Escherichia coli. Here, we show that DNA supercoiling alleviates the observed cold-sensitivity of promoter opening by the Deinococcus-Thermus RNAPs and by the hybrid holoenzymes and allows melting of the transcription start site at the same temperatures as in the case of E. coli RNAP. Supercoiling also suppresses salt sensitivity of the promoter complexes formed by these RNAPs. The results demonstrate that the RNAPs from Deinococcus-Thermus species are sensitive to DNA supercoiling and suggest that they can be rapidly switched-off or activated by the supercoiling state of the host genomes.

Keywords: DNA supercoiling; Deinococcus radiodurans; RNA polymerase; promoter melting; stress resistance.