Semester

Summer

Date of Graduation

2005

Document Type

Dissertation

Degree Type

PhD

College

School of Medicine

Department

Microbiology, Immunology, and Cell Biology

Committee Chair

Thomas Elliott.

Abstract

In enteric bacteria, stress adaptation is mediated by the RpoS protein, one of several sigma-factors that, in association with RNA polymerase, collectively allow a tailored transcriptional response to environmental cues. Stress stimuli including low temperature, osmotic shock, and starvation all result in a substantial increase in RpoS abundance. Perhaps the most pronounced affect is observed during growth to stationary phase (SP) in rich medium. The mechanism of regulation depends on the specific signal, but may occur at the level of transcription, translation, protein activity or targeted proteolysis. In both Escherichia coli and Salmonella enterica cultured in rich undefined medium, the RpoS protein is barely detectable during exponential growth and increases >30-fold as cells enter SP. Under these conditions, SP induction depends on transcriptional and translational control with proteolysis affecting basal levels but not regulation per se. The transiently expressed Fis protein, whose abundance inversely correlates to that of RpoS, binds just upstream of the primary rpoS promoter and represses transcription nearly 10-fold specifically during exponential growth. SP induction at the translational level relies on a novel form of genetic control dependent on the 24 nucleotides preceding the rpoS initiation codon (ribosome-binding sequence, RBS). The RNA secondary structure of the rpoS RBS is necessary and sufficient for a nearly 10-fold translational increase during SP. Control at this level is not a result of differential transcript stability, nor does it involve the known rpoS regulators ppGpp, DksA, HU, Hfq or the small regulatory RNAs, DsrA and RprA. The environmental stimuli that trigger RBS-mediated SP induction of rpoS translation also remain unknown, but similar to transcriptional control, regulation is only seen in rich undefined media. Collectively, transcriptional repression by Fis and RBS-mediated induction at the translational level account for approximately 95% of the overall SP induction of RpoS.

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