Genome-wide studies of the regulon members of alternative sigma factors, sigma32 and sigma28, in Escherichia coli

Escherichia coli DNA-dependent RNA polymerase is a central enzyme in the cell and exists in two forms. Core enzyme is required for transcription of DNA, yet alone the enzyme is unable to initiate promoter specific transcription. RNA polymerase holoenzyme which consists of core enzyme and an additional subunit, the sigma factor, allows recognition of specific promoter sequences and confers the ability to regulate global gene expression. Sigma regulons studies should be able to provide valuable information for determining functions of individual genes as well as for comprehensively understanding E. coli physiology. In my project, transcriptional responses of E. coli to the concentration changes of two alternative sigma factors, sigma 32 (heat-shock regulation) and sigma28 (flagellar-motility regulation), were studied.; The global transcriptional response of E. coli to induced sigma 32/sigmaF protein allow us to successfully confirm genes previous known to be directly under the control of sigma32 /sigmaF and also to assign many additional genes to sigma32/sigmaF regulon. Measuring both the protein level of sigma32 and the transcriptional levels of sigma32-controlled genes led us to characterize gene expression dynamics as a function of time after sigma32 induction. We discovered that a decrease in sigma32 activity rather than in sigma 32 level is responsible for the rapid shutoff of the transcription of sigma32-dependent genes in our assays. Further experiments suggest DnaK can act as an anti-sigma factor to functionally inactivate sigma 32 and thus reduce sigma32-dependent transcription in vivo. In sigmaF regulon studies, we also characterize "foraging"-like behaviors in E. coli with response to different carbon sources. The synchronized pattern of increasing CRP activity causing increasing FlhDC transcription with decreasing carbon source quality, and the apparent coupling of motility activity and activation of motility and chemotaxis genes in poor quality carbon sources. These results highlight the important role of CRP activation in bacteria risk-prone foraging behaviors which can potentially increase a cell's ability for maximizing survival by strategically using the precious energy in nutrient-poor environments.