| Abstract: | Xanthomonas campestris pv. campestris (Xcc) is the pathogen causing black rot in cruciferous plants. It has previously demonstrated in our laboratory that a fleQ mutant is non-flagellated and immobile and expression of flagella genes fliE, fliQ, fliL, flgG, flgB, and flhF was depended on transcriptional activator, FleQ.
In this work, it is shown that expression of fliA and fliC is also regulated by FleQ. In order to know if a plasmid-encoded fleQ gene can complement a mutation in fleQ, the fleQ gene was cloned into pBBad22K carrying an arabinose inducible promoter. The derived plasmid, pBadfleQ, was introduced into mutants and the parental strain. The phenotypic and Western blot analysis demonstrated that plasmid-encoded fleQ was expressed in the absence of inducer and the small amount of FleQ produced was sufficient to complement the mobility defect of a fleQ mutant. Addition of arabinose was increased the production of FleQ but decreased the effect of complementation. Induction to overexpress FleQ by 0.05% arabinose in Xc17fleQ::Gm (pBadfleQ ) and Xc17(pBadfleQ ) hindered their growth and severely reduced the biosynthesis of extracellular polysaccharide (EPS) and extracellular enzymes, such as amylases, proteinases, and pectinases. The promoter activity of the major EPS biosynthesis gene gumB is inhibited by the overexpressed FleQ protein. Since EPS and extracellular enzymes are important virulence factors in Xcc, Xc17fleQ::Gm(pBadfleQ) and Xc17(pBadfleQ), are less virulent than their parental wild-Type strain.
It has previously been show in our laboratory that RpoN, FleQ, FleN, FlhF, FlgM and FliA can regulate flagellar genes expression. In this work, yeast two-hybrid system was used to further analyze the protein-protein interaction between these regulators. Our data revealed that proteins FleN and FliA and FlgM and FliA, RpoN2 and FlhF have interaction. |
