The E. coli Anti-Sigma Factor Rsd: Studies on the Specificity and Regulation of Its Expression

Background Among the seven different sigma factors in E. coli σ 70 has the highest concentration and affinity for the core RNA polymerase. The E. coli protein Rsd is regarded as an anti-sigma factor, inhibiting σ 70 -dependent transcription at the onset of stationary growth. Although binding of Rsd to σ 70 has been shown and numerous structural studies on Rsd have been performed the detailed mechanism of action is still unknown. Methodology/Principal Findings We have performed studies to unravel the function and regulation of Rsd expression in vitro and in vivo . Cross-linking and affinity binding revealed that Rsd is able to interact with σ 70 , with the core enzyme of RNA polymerase and is able to form dimers in solution. Unexpectedly, we find that Rsd does also interact with σ 38 , the stationary phase-specific sigma factor. This interaction was further corroborated by gel retardation and footprinting studies with different promoter fragments and σ 38 - or σ 70 -containing RNA polymerase in presence of Rsd. Under competitive in vitro transcription conditions, in presence of both sigma factors, a selective inhibition of σ 70 -dependent transcription was prevailing, however. Analysis of rsd expression revealed that the nucleoid-associated proteins H-NS and FIS, StpA and LRP bind to the regulatory region of the rsd promoters. Furthermore, the major promoter P2 was shown to be down-regulated in vivo by RpoS, the stationary phase-specific sigma factor and the transcription factor DksA, while induction of the stringent control enhanced rsd promoter activity. Most notably, the dam -dependent methylation of a cluster of GATC sites turned out to be important for efficient rsd transcription. Conclusions/Significance The results contribute to a better understanding of the intricate mechanism of Rsd-mediated sigma factor specificity changes during stationary phase.