Identification of transcriptional start sites and the role of ppGpp in the expression of rpoS, the structural gene for the sigma S subunit of RNA polymerase in Escherichia coli

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Abstract
RpoS is the structural gene for the sigma S subunit of RNA polymerase which controls the expression of a large number of genes in Escherichia coli that are induced during entry into stationary phase or in response to increased medium osmolarity. Using a combination of primer extension experiments and a 59 deletion analysis of the region upstream of rpoS, we show that rpoS transcription is mainly driven by a single promoter (rpoSp1) located within the nlpD gene upstream of rpoS (the two relatively weak nlpD promoters contribute to the low level of rpoS expression during early exponential phase). In addition, we demonstrate that the expression of both transcriptional and translational rpoS::lacZ fusions as well as the level of rpoS mRNA originating at rpoSp1 is strongly reduced in ppGpp-deficient relA spoT mutants. However, experiments with the 59 deletion constructs indicate that a lack of ppGpp does affect transcriptional elongation rather than initiation.