Transcription‐coupled repair (TCR) of damaged DNA in E. coli can involve several different pathways. We propose that one pathway entails release of RNAP at the site of DNA damage without dissociation of the RNA:DNA hybrid. The RNA can serve as a primer for PolI, which initiates the repair process. We are investigating how TCR repairs nalidixic acid‐induced DNA damage. Nalidixic acid, like Etoposide generates a DNA double‐strand break bearing a TopA adduct at each 5′ DNA end. We show that efficient repair requires two transcription factors. The first DksA, is a coiled‐coil protein that inserts into the RNAP secondary channel and, in concert with ppGpp, regulates the activity of certain promoters. The role of DksA in transcription elongation is undefined, although it is known to suppresses replisome – RNAP clashes. DksA plays a direct role in repair, rather than blocking access of GreA to RNAP. Thus, a ΔdksA mutation is not suppressed by a greA deletion. Recent work from the Murakami laboratory reveals that DksA binding distorts the structure of RNAP, opening the pincers, thus potentially reducing the processivity of transcription elongation. DksA in association with ppGpp, however, does not affect RNAP structure. We find that repair is more efficient in strains carrying an RNAP mutation that reduces ppGpp binding to site 2, the location of DksA insertion. We have isolated a suppressor of ΔdksA based on resistance to nalidixic acid. The mutation, rpoBQ148H, phenocopies RNAP bound to DksA in the absence of ppGpp, i.e. it opens the pincers and may favor release of elongating RNAP. These findings suggest that DksA may act as a transcription termination factor that can release RNAP without removing the RNA:DNA hybrid.The second, transcription termination protein Rho, is an RNA‐dependent DNA:RNA helicase. The defects in TCR of rho and dksA mutations are additive, suggesting that they act on different substrates. Whereas Rho is thought to release RNAP by unwinding the RNA:DNA helix, rho mutants that terminate transcription without releasing RNA have been isolated.Another reaction that entails elongation of an RNA:DNA hybrid by PolI is constitutive stable DNA replication (cSDR). In cSDR, elongation of an R‐loop by PolI creates a new origin of replication. We find that cSDR is dependent on DksA. Presumably the mechanisms of nalidixic acid DNA damage and cSDR both involve allowing access of PolI to the 3′ end of an RNA:DNA hybrid.Support or Funding InformationGM037219 GM037219 NIH: GM037219 NIH: GM037219 GM037219This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.