• Characterization of Novel Factors Involved in Swimming and Swarming Motility in Salmonella enterica Serovar Typhimurium.

      Deditius, Julia Andrea; Felgner, Sebastian; Spöring, Imke; Kühne, Caroline; Frahm, Michael; Rohde, Manfred; Weiß, Siegfried; Erhardt, Marc; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig, Germany. (2015)
      Salmonella enterica utilizes flagellar motility to swim through liquid environments and on surfaces. The biosynthesis of the flagellum is regulated on various levels, including transcriptional and posttranscriptional mechanisms. Here, we investigated the motility phenotype of 24 selected single gene deletions that were previously described to display swimming and swarming motility effects. Mutations in flgE, fliH, ydiV, rfaG, yjcC, STM1267 and STM3363 showed an altered motility phenotype. Deletions of flgE and fliH displayed a non-motile phenotype in both swimming and swarming motility assays as expected. The deletions of STM1267, STM3363, ydiV, rfaG and yjcC were further analyzed in detail for flagellar and fimbrial gene expression and filament formation. A ΔydiV mutant showed increased swimming motility, but a decrease in swarming motility, which coincided with derepression of curli fimbriae. A deletion of yjcC, encoding for an EAL domain-containing protein, increased swimming motility independent on flagellar gene expression. A ΔSTM1267 mutant displayed a hypermotile phenotype on swarm agar plates and was found to have increased numbers of flagella. In contrast, a knockout of STM3363 did also display an increase in swarming motility, but did not alter flagella numbers. Finally, a deletion of the LPS biosynthesis-related protein RfaG reduced swimming and swarming motility, associated with a decrease in transcription from flagellar class II and class III promoters and a lack of flagellar filaments.
    • RflM mediates target specificity of the RcsCDB phosphorelay system for transcriptional repression of flagellar synthesis in Salmonella enterica.

      Kühne, Caroline; Singer, Hanna M; Grabisch, Eva; Codutti, Luca; Carlomagno, Teresa; Scrima, Andrea; Erhardt, Marc; Helmholtz Centre for infection research, Inhoffenstr. 7, 38124 Braunschweig. (2016-09)
      The bacterial flagellum enables directed movement of Salmonella enterica towards favorable conditions in liquid environments. Regulation of flagellar synthesis is tightly controlled by various environmental signals at transcriptional and post-transcriptional levels. The flagellar master regulator FlhD4 C2 resides on top of the flagellar transcriptional hierarchy and is under autogenous control by FlhD4 C2 -dependent activation of the repressor rflM. The inhibitory activity of RflM depends on the presence of RcsB, the response regulator of the RcsCDB phosphorelay system. In this study, we elucidated the molecular mechanism of RflM-dependent repression of flhDC. We show that RcsB and RflM form a heterodimer that coordinately represses flhDC transcription independent of RcsB phosphorylation. RcsB-RflM complex binds to a RcsB box downstream the P1 transcriptional start site of the flhDC promoter with increased affinity compared to RcsB in the absence of RflM. We propose that RflM stabilizes binding of unphosphorylated RcsB to the flhDC promoter in absence of environmental cues. Thus, RflM is a novel auxiliary regulatory protein that mediates target specificity of RcsB for flhDC repression. The cooperative action of the RcsB-RflM repressor complex allows Salmonella to fine-tune initiation of flagellar gene expression and adds another level to the complex regulation of flagellar synthesis.