FliZ Is a posttranslational activator of FlhD4C2-dependent flagellar gene expression

Flagellar assembly proceeds in a sequential manner, beginning at the base and concluding with the filament. A critical aspect of assembly is that gene expression is coupled to assembly. When cells transition from a nonflagellated to a flagellated state, gene expression is sequential, reflecting the manner in which the flagellum is made. A key mechanism for establishing this temporal hierarchy is the sigma(28)-FlgM checkpoint, which couples the expression of late flagellar (P(class3)) genes to the completion of the hook-basal body. In this work, we investigated the role of FliZ in coupling middle flagellar (P(class2)) gene expression to assembly in Salmonella enterica serovar Typhimurium. We demonstrate that FliZ is an FlhD(4)C(2)-dependent activator of P(class2)/middle gene expression. Our results suggest that FliZ regulates the concentration of FlhD(4)C(2) posttranslationally. We also demonstrate that FliZ functions independently of the flagellum-specific sigma factor sigma(28) and the filament-cap chaperone/FlhD(4)C(2) inhibitor FliT. Furthermore, we show that the previously described ability of sigma(28) to activate P(class2)/middle gene expression is, in fact, due to FliZ, as both are expressed from the same overlapping P(class2) and P(class3) promoters at the fliAZY locus. We conclude by discussing the role of FliZ regulation with respect to flagellar biosynthesis based on our characterization of gene expression and FliZ's role in swimming and swarming motility.