Analysis of a putative RNA repair operon in Salmonella Typhimurium
Hartman, Christine Elizabeth
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The genome of Salmonella Typhimurium encodes a putative RNA repair operon, comprising three structural genes: rsr, rtcB, and rtcA; and two non-coding RNAs: yrlB and yrlA. Rsr is involved in non-coding RNA quality control, forming a ribonucleoprotein complex with YrlA and PNPase to degrade misfolded structured RNAs. RtcB has both DNA and RNA ligase activity and can “cap” 3′ phosphate ends of nicked double stranded DNA to protect the end from exonuclease activity. RtcA repairs damaged ends of RNA by converting 2′ or 3′ phosphate ends to 2′,3′ cyclic phosphates, which can serve as a substrate for ligation by RtcB. Transcription of the RNA repair operon is controlled by the highly regulated sigma factor, σ54, and the bacterial enhancer binding protein, RtcR. To activate transcription, RtcR recognizes an as yet unknown signal from the cell to become active, multimerize, and bind an enhancer sequence upstream of the RNA repair operon. ATP hydrolysis by RtcR is then required for open complex formation and transcription initiation. To date, no published work has shown RtcR-dependent activation of the RNA repair operon. To determine the cellular signal that induces transcription, we treated Salmonella Typhimurium cultures with stressors known to damage nucleic acids and therefore may produce the damaged substrates on which the components of the RNA repair system act. Treatment with Mitomycin C, hydrogen peroxide, and nitrogen limitation resulted in increased expression of the RNA repair operon. Mitomycin C produced the greatest and only RtcR-dependent activation of the operon, and was further studied for characterization of the RNA repair system through viability assays, RNA-seq analysis, and investigations into potential interacting partners of the components of the system.