Abstract
The carbon storage regulator protein CsrA regulates cellular processes post-transcriptionally by binding to target-RNAs altering translation efficiency and/or their stability. Here we identified and analyzed the direct targets of CsrA in the human pathogen Legionella pneumophila. Genome wide transcriptome, proteome and RNA co-immunoprecipitation followed by deep sequencing of a wild type and a csrA mutant strain identified 479 RNAs with potential CsrA interaction sites located in the untranslated and/or coding regions of mRNAs or of known non-coding sRNAs. Further analyses revealed that CsrA exhibits a dual regulatory role in virulence as it affects the expression of the regulators FleQ, LqsR, LetE and RpoS but it also directly regulates the timely expression of over 40 Dot/Icm substrates. CsrA controls its own expression and the stringent response through a regulatory feedback loop as evidenced by its binding to RelA-mRNA and links it to quorum sensing and motility. CsrA is a central player in the carbon, amino acid, fatty acid metabolism and energy transfer and directly affects the biosynthesis of cofactors, vitamins and secondary metabolites. We describe the first L. pneumophila riboswitch, a thiamine pyrophosphate riboswitch whose regulatory impact is fine-tuned by CsrA, and identified a unique regulatory mode of CsrA, the active stabilization of RNA anti-terminator conformations inside a coding sequence preventing Rho-dependent termination of the gap operon through transcriptional polarity effects. This allows L. pneumophila to regulate the pentose phosphate pathway and the glycolysis combined or individually although they share genes in a single operon. Thus the L. pneumophila genome has evolved to acclimate at least five different modes of regulation by CsrA giving it a truly unique position in its life cycle.
Highlights
The Gram negative, environmental bacterium Legionella pneumophila is proliferating in aquatic environments where it parasitizes in fresh water protozoa [1,2,3]
Proteomics, RNA-Immunoprecipitation followed by deep sequencing (RIPseq), together with biochemical, phenotypical and molecular analyses we identified the L. pneumophila CsrA targets genome wide and discovered a new mode of action of CsrA that allows to regulate genes comprised in the same operon, independently
CsrA is essential for L. pneumophila, but such a truncated CsrA variant has a strongly reduced expression of CsrA (S1B Fig), to what was reported for Escherichia coli [26, 27], possibly by immediate degradation of a miss folded protein due to the distorted C-terminal helical structure
Summary
The Gram negative, environmental bacterium Legionella pneumophila is proliferating in aquatic environments where it parasitizes in fresh water protozoa [1,2,3]. The current model of the L. pneumophila life cycle regulation is that starvation of amino acids and altered fatty acid biosynthesis lead to the production of (p)ppGpp and subsequently the activation of the two-component system (TCS) LetA/LetS and the alternative sigma factor RpoS [14, 15]. Both promote the transcription of the small non-coding RNAs RsmX, RsmY and RsmZ, which in turn bind and sequester CsrA leading to the expression of transmissive and repression of replicative traits [16,17,18]. In Legionella, PmrBA was shown to regulate the expression of several Dot/Icm effector proteins and positively regulates the transcription of csrA [16, 24]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
