Abstract
In Escherichia coli, physical collisions between the replication machinery and transcription complexes are an unavoidable consequence of DNA replication and transcription happening simultaneously. These conflicts are exacerbated by DNA damage, which stalls replication forks and can trap RNA polymerase (RNAP) on damaged DNA templates. While the canonical response to DNA damage is the LexA/RecA-mediated SOS regulon, evidence supports the existence of parallel, SOS-independent pathways that sense replication stress through the activity state of the replication initiator protein DnaA. This thesis investigates the transcriptional regulation of rapA, which encodes an RNAP-associated ATPase that promotes dissociation of post-termination complexes (PTCs), and its relationship to the stress response factor SspA under conditions of ciprofloxacin (CPX) induced replication stress.Using luciferase-based assays, we show that the rapA promoter is induced by CPX in a manner independent of the SOS response, and that rapA expression is under repression by DnaA in its ATP-bound form. Additionally, rapA induction is consistently elevated in a sspAΔ background, suggesting that SspA modulates rapA expression either directly through transcriptional regulation or indirectly through its competitive occupancy of the same RNAP binding site. Together, these results support a model in which replication stress relieves DnaA-mediated repression of rapA, and in which the competition for RNAP binding of RapA and SspA determines the efficiency of PTC clearance during the DNA damage response.