The Importance of a Fifth Element in Transcription: Transcriptional Discrimination in Escherichia coli

Upon growth arrest of the bacterium Escherichia coli, RNA polymerase (Es70) is redirected from transcribing genes encoding the protein synthesizing system (PSS) to those involved in maintenance and stress resistance. The small nucleotide ppGpp, which is directly targeting Es70, is a key regulatory molecule required for this response. In addition, the protein DksA has been hypothesized to be required for the regulatory function of ppGpp. Cells defective in the synthesis of either ppGpp or DksA do not undergo a shift in gene expression upon growth arrest and are deficient in maintenance-related activities. The underlying mechanism of how regulation of gene expression is exerted by ppGpp and DksA is the main focus of this thesis. -Text removed from public version- On top of a direct role of ppGpp in regulating gene expression, ppGpp can potentially affect gene regulation passively by modulating the levels of free Es70 in the cell. Using a mini-cell approach, I demonstrate an inverse correlation between the levels of ppGpp and free Es70. Thus, I hypothesize that ppGpp contribute to the redistribution of Es70 at promoters during growth arrest also by decreasing free Es70 levels, which would negatively affect promoters requiring high levels of Es70 for efficient transcription, such as those of the PSS