Structure of the periplasmic domain of SflA involved in spatial regulation of the flagellar biogenesis of Vibrio reveals a TPR/SLR-like fold.

Bacteria have evolved various types of flagellum, an organella for bacterial motility, to adapt to their habitat environments. The number and the spatial arrangement of the flagellum are precisely controlled to optimize performance of each type of the flagellar system. Vibrio alginolyticus has a single sheathed flagellum at the cell pole for swimming. SflA is a regulator protein to prevent peritrichous formation of the sheathed flagellum, and consists of an N-terminal periplasmic region, a transmembrane helix, and a C-terminal cytoplasmic region. Whereas the cytoplasmic region has been characterized to be essential for inhibition of the peritrichous growth, the role of the N-terminal region is still unclear. We here determined the structure of the N-terminal periplasmic region of SflA (SflAN) at 1.9-Å resolution. The core of SflAN forms a domain-swapped dimer with tetratricopeptide repeat (TPR)/Sel1-like repeat (SLR) motif, which is often found in the domains responsible for protein-protein interaction in various proteins. The structural similarity and the following mutational analysis based on the structure suggest that SflA binds to unknown partner protein by SflAN and the binding signal is important for the precise control of the SflA function.

Localization and domain characterization of the SflA regulator of flagellar formation in Vibrio alginolyticus.

Many swimming bacteria use flagella as locomotive organelles. The spatial and numerical regulation of flagellar biosynthesis differs by bacterial species. The marine bacteria Vibrio alginolyticus use a single polar flagellum whose number is regulated positively by FlhF and negatively by FlhG. Cells lacking FlhF and FlhG have no flagellum. The motility defect in an flhFG deletion was suppressed by a mutation in the sflA gene that resulted in the production of multiple, peritrichous flagella. SflA is a Vibrio-specific protein. SlfA either facilitates flagellum growth at the cell pole or prevents flagellar formation on the cell body by an unknown mechanism. Fluorescent protein fusions to SflA localized to the cell pole in the presence of FlhF and FlhG, but exhibited both polar and lateral cell localization in ΔflhFG cells. Polar localization of SflA required the polar landmark protein HubP. Over-expression of the C-terminal region of SflA (SflAC ) in ΔflhFG ΔsflA cells suppressed the lateral flagellar formation. Our results suggest that SflA localizes with the flagella and that SflAC represses the flagellar initiation in ΔflhFG strains. A model is presented where SflA inhibits lateral flagellar formation to facilitate single polar flagellum assembly in V. alginolyticus cells.

A novel dnaJ family gene, sflA, encodes an inhibitor of flagellation in marine Vibrio species.

The marine bacterium Vibrio alginolyticus has a single polar flagellum. Formation of that flagellum is regulated positively and negatively by FlhF and by FlhG, respectively. The ΔflhF mutant makes no flagellum, whereas the ΔflhFG double-deletion mutant usually lacks a flagellum. However, the ΔflhFG mutant occasionally reverts to become motile by forming peritrichous flagella. We have isolated a suppressor pseudorevertant from the ΔflhFG strain (ΔflhFG-sup). The suppressor strain forms peritrichous flagella in the majority of cells. We identified candidate suppressor mutations by comparing the genome sequence of the parental strain, VIO5, with the genome sequences of the suppressor strains. Two mutations were mapped to a gene, named sflA (suppressor of ΔflhFG), at the VEA003730 locus of the Vibrio sp. strain EX25 genome. This gene is specific for Vibrio species and is predicted to encode a transmembrane protein with a DnaJ domain. When the wild-type gene was introduced into the suppressor strain, motility was impaired. Introducing a mutant version of the sflA gene into the ΔflhFG strain conferred the suppressor phenotype. Thus, we conclude that loss of the sflA gene is responsible for the suppressor phenotype and that the wild-type SflA protein plays a role in preventing polar-type flagella from forming on the lateral cell wall.