A suppressor mutation approach to determining the sequence requirements of an unusual two-subunit sigma factor and its cognate promoter element

Abstract

Transcription is a highly-regulated process. In bacteria, σ factors are transcription factors that regulate RNA polymerase binding to promoters, and the conversion of double stranded DNA to the single-stranded state required for the initiation of RNA synthesis. The Bacillus subtilis genome encodes an atypical, twosubunit σ factor called SigO-RsoA. To better understand the genes regulated by this σ factor, I conducted the largest phylogenetic analysis of the SigO-RsoA regulon to date and compiled and analyzed the sequences of 216 orthologous target promoters. A comprehensive mutational analysis of a conserved thymine trinucleotide and a -10 promoter element cognate to SigO-RsoA delineated key nucleotides required for the activity of this σ factor. Selected inactive promoter mutants were used in a suppressor mutation approach to identify whether it is SigO or RsoA that is responsible for interaction with the key -10 promoter element.