Sequence Determinants Spanning -10 Motif and Spacer Region Implicated in Unique Sigma 32-Dependent Promoter Activity of Gene.

is an obligate intracellular tick-borne bacterium that causes human monocytic ehrlichiosis. Studying gene regulation is challenge, as this and related rickettsiales lack natural plasmids and mutagenesis experiments are of a limited scope. contains only two sigma factors, σ and σ. We previously developed surrogate system to study transcriptional regulation from RNA polymerase (RNAP) containing σ or σ. We reported that RNAP binding motifs of genes recognized by σ or σ share extensive homology and that transcription may be initiated by either one of the sigma factors, although transcriptional efficiencies differ. In the current study, we investigated mapping the gene promoter using the pathogen σ expressed in lacking its native σ. The surrogate system and our previously described transcription system aided in defining the unique -10 motif and spacer sequence of the promoter. We also mapped σ amino acids/domains engaged in its promoter regulation in . The data reported in this study demonstrate that the -10 and -35 motifs and spacer sequence located between the two motifs of promoter are critical for the RNAP function. Further, we mapped the importance of all six nucleotide positions of the -10 motif and identified critical determinants within it. In addition, we reported that the lack of C-rich sequence upstream to the -10 motif is unique in driving the pathogen-specific transcription by its σ from gene promoter. This is the first study in defining an σ-dependent promoter and it offers insights about how this and other related rickettsial pathogens regulate stress response genes.