| The bacterial flagellum is one of the most remarkable nanomachine that allows cells to move in their environment. So, it has profound significance to study its assembly and regulation for understanding the molecular mechanisms of bacterial environmental adaptability. As a most conserved molecular machine made up of a large number of structural subunits, the flagellum is under tight regulation by hierarchical arrangements. Although variations in polar flagellar systems are found, they are largely restricted to multiple-copy components, such as flagellins and stators. Therefore, this feature is regarded to be peripheral relative to the comprehensive conservation. In this study, however, we present evidence to show that the difference in highly conserved polar flagellar systems can be surprisingly profound, even at the heart of the classical regulatory hierarchy.Regulation of FlrBC on flagellar biosynthesis and flagellin expression in Shewanella oneidensisIn Gram-negative S. oneidensis, two-component system (TCS) FlrBC, whose counterparts studied thus far are essential for flagellar biosynthesis and motility by directly controlling expression of class III genes in polarly flagellated bacteria such as Vibrio cholerae, is dispensable for these processes. In contrast, loss of both flrB and flrC results in a hypermotile phenotype although the TCS is still regulated by σ54 and the top-tier regulator FlrA and affects expression of the downstream genes, which are not restricted to the class III of the typical regulatory hierarchy. We further show that the ratio of two flagellins, FlaA and FlaB, finally determines motility of a flagellum and the FlrBC-associated hypermotility results from overproduction of FlaB.Roles of FlrC in flagellar polarization and number control, as well as the other aspects of S. oneidensisOverproduction of FlrC results in a peritrichously multi-flagellated phenotype. This is not caused by the reduced expression of flhG, which encodes a protein functioning in the localization of flagella in a variety of bacteria. FlrC directly controls expression of the flaA gene and is likely to act as an activator in its unphosphorylated form, contrasting to the previously characterized counterparts. Further we find that deletion of gene SO4002 encoding a sensor kinase shows an additive effect on hypermotility of the flrBC mutant by means of the transposon mutagenesis screening. Thus, SO4002 may also phosphorylate regulator FlrC. In addition, loss of the TCS influences conjunctive efficiency of the receptor strain.Exoproteins correlate with morphotype changes of nonmotile aflagellate S. oneidensis mutants.We report a previously undescribed mechanism for the rugose morphotype in S. oneidensis. Bacteria may form smooth or rugose colonies on agar plates. In general, conversion from the smooth to rugose colony morphotype is attributed to increased production of exopolysaccharide (EPS). We discover that aflagellate S. oneidensis mutants grow into rugose colonies, whereas those with nonfunctional flagellar filaments remain smooth. EPS production is not altered in either case, but mutants with the rugose morphotype show significantly reduced exoproteins. The idea that exoproteins at a reduced level correlate with rugosity gains support from smooth suppressor strains of an aflagellate rugose JliD (encoding the capping protein) mutant, which restore the exoprotein level to the levels of the wild-type and mutant strains with a smooth morphotype. Further analyses reveal that SO1072 (a putative GlcNAc-binding protein) is one of the highly upregulated exoproteins in these suppressor strains. Most intriguingly, this study identifies a compensatory mechanism of SO1072 to flagellins possibly mediated by bis-(3’-5’)-cyclic dimeric GMP. |
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