Analysis of Potential Transcriptional Regulators of the Universal Stress Protein A (UspA) Gene of Escherchia coli

The universal stress protein A (UspA) of E. coli is induced by a wide variety of stressful conditions and appears to be important in regulating the cell\u27s overall stress response. However, the exact mechanism of uspA induction remains unknown. The purpose of this project was to identify potential regulators of the uspA gene. Regulators tested included the LexA repressor of the SOS regulon, and the sigma factors RpoH and RpoS. Sigma factors initiate transcription by recognizing target promoters and allowing RNA polymerase to bind. Most sigma factors are involved in allowing the cell to respond to a particular stressful condition, such as membrane stress or nutrient starvation. Using a bioluminescent reporter system in which the uspA promoter is fused to the lux operon from Vibrio fischeri, the level of uspA transcription under different stress conditions was quantified. When stressed with 5 mJ/m² UV radiation, lexA-independent mutants showed, on average, a six-fold increase in uspA induction compared to the wildtype strain. The lexA-independent strain encodes a mutant form of the LexA repressor that cannot be cleaved by RecA and thus remains bound to the promoters of genes in the SOS regulon. The increased transcription levels of uspA in the lexA-independent strain suggest that uspA is not a member of the SOS regulon. Similarly, when ethanol stressed, uspA transcription in the rpoH mutant strain showed a four-fold increase over the rpoH wildtype strain, suggesting that RpoH is not involved in initiating uspA transcription. Conversely, uspA transcription in rpoS mutants was reduced by almost 70% during stationary phase compared to the rpoS wildtype strain. This suggests that either the RpoS protein itself or the product of an RpoS-upregulated gene is important in initiating uspA transcription