Journal of Bacteriology
Genomics, Non-coding RNA, Bacterial genomes
Coxiella burnetii is an obligate intracellular gammaproteobacterium and zoonotic agent of Q fever. We previously identified 15 small non-coding RNAs (sRNAs) of C. burnetii. One of them, CbsR12 (Coxiella b urnetii small RNA 12), is highly transcribed during axenic growth and becomes more prominent during infection of cultured mammalian cells. Secondary structure predictions of CbsR12 revealed four putative CsrA-binding sites in stem loops with consensus AGGA/ANGGA motifs. We subsequently determined that CbsR12 binds to recombinant C. burnetii CsrA-2, but not CsrA-1, proteins in vitro. Moreover, through a combination of in vitro and cell culture assays, we identified several in-trans mRNA targets of CbsR12. Of these, we determined that CbsR12 binds and upregulates translation of carA transcripts coding for carbamoyl phosphate synthetase A; an enzyme that catalyzes the first step of pyrimidine biosynthesis. In addition, CbsR12 binds and downregulates translation of metK transcripts coding for S-adenosyl methionine (SAM) synthetase, a component of the methionine cycle. Furthermore, we found that CbsR12 binds to and downregulates the quantity of cvpD transcripts, coding for a type IVB effector protein, in mammalian cell culture. Finally, we found that CbsR12 is necessary for expansion of Coxiella-containing vacuoles (CCVs) and affects growth rates in a dose-dependent manner in the early phase of infecting THP-1 cells. This is the first characterization of a trans-acting sRNA of C. burnetii and first example of a bacterial sRNA that regulates both CarA and MetK synthesis. CbsR12 is one of only a few identified trans-acting sRNAs that interacts with CsrA.
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Published as: Wachter, S., Bonazzi, M., Shifflett, K., Moses, A. S., Raghavan, R., & Minnick, M. F. (2019). A CsrA-binding, trans-acting sRNA of Coxiella burnetii is necessary for optimal intracellular growth and vacuole formation during early infection of host cells. Journal of bacteriology, JB-00524.