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Binding interface between the Salmonella σ(S)/RpoS subunit of RNA polymerase and Crl: hints from bacterial species lacking crl.

Abstract : In many Gram-negative bacteria, including Salmonella enterica serovar Typhimurium (S. Typhimurium), the sigma factor RpoS/σ S accumulates during stationary phase of growth, and associates with the core RNA polymerase enzyme (E) to promote transcription initiation of genes involved in general stress resistance and starvation survival. Whereas σ factors are usually inactivated upon interaction with anti-σ proteins, σ S binding to the Crl protein increases σ S activity by favouring its association to E. Taking advantage of evolution of the σ S sequence in bacterial species that do not contain a crl gene, like Pseudomonas aeruginosa, we identified and assigned a critical arginine residue in σ S to the S. Typhimurium σ S-Crl binding interface. We solved the solution structure of S. Typhimurium Crl by NMR and used it for NMR binding assays with σ S and to generate in silico models of the σ S-Crl complex constrained by mutational analysis. The σ S-Crl models suggest that the identified arginine in σ S interacts with an aspartate of Crl that is required for σ S binding and is located inside a cavity enclosed by flexible loops, which also contribute to the interface. This study provides the basis for further structural investigation of the σ S-Crl complex. In bacteria, a primary housekeeping sigma factor and one or more alternative σ factors associate with the catalytically active RNA polymerase (RNAP) core enzyme (α 2β β 'ω , E), to form the holoenzyme Eσ , and direct transcription initiation of specific subsets of genes 1,2. In many Gram-negative bacteria, σ S /RpoS is produced during late exponential phase, or in response to stress, to modify global gene transcription and to allow stationary phase survival and general stress resistance 3–5. In the wide host-range pathogen S. Typhimurium, σ S is not only required for general stress resistance, but also for virulence, biofilm formation and development of the red dry and rough (rdar) morphotype, a colony morphology caused by the production of amyloid fibers (curli) and cellulose 6–8. The efficiency of formation of the housekeeping and alternative Eσ can be modulated by regulatory factors that bind E and/or σ 5,9. So far, Crl is the only known σ S-dedicated regulatory factor that enhances σ S activity through a direct interaction, favouring Eσ S formation 7,10–15. Analyses of sequenced bacterial genomes revealed that crl is less widespread and less conserved at the sequence level than
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Paola Cavaliere, Christina Sizun, Fabienne Levi-Acobas, Mireille Nowakowski, Véronique Monteil, et al.. Binding interface between the Salmonella σ(S)/RpoS subunit of RNA polymerase and Crl: hints from bacterial species lacking crl.. Scientific Reports, Nature Publishing Group, 2015, 5, pp.13564. ⟨10.1038/srep13564⟩. ⟨pasteur-01392714⟩

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