Chapter 16 : Regulation of Flagellum Biosynthesis and Motility in Caulobacter

C. crescentus cell division cycle and temporal expression pattern of flagellar genes. The chromosome is represented by an oval, with the "theta" structure indicating DNA replication. The times during which DNA synthesis occurs and the major classes of flagellar genes expressed are delineated beneath the relevant structural changes taking place during the cell cycle. The expression of the single known class I gene, ctrA, is apparently initiated concurrently with DNA replication.

Structure of the C. crescentus flagellum. In this schematic illustration, the components that make up the flagellum are indicated with respect to position in the membrane, shown in cross section. The names o f the genes that encode the relevent structural proteins are shown in parentheses below each component name. The individual components are grouped into three major superstructures: the basal body, the hook, and the filament. The basal body includes the cytoplasmic structures and the structures that span the inner (cytoplasmic) membrane (IM), the peptidoglycan layer (PL) in the periplasmic space, and the outer membrane (OM) of the cell. Outside the cell, the flexible hook connects the basal body to the rigid flagellar filament.

Assembly of the C. crescentus flagellum. The upper diagram shows the flagellum at the major stages of cell-proximal-to-cell-distal assembly. Below, the regulatory hierarchy is outlined, with the genes expressed at each step in the cascade shown in boxes beneath the corresponding structures. The roles of specific proteins controlling expression of each gene class are illustrated with arrows for positive regulation and lines with bars for negative regulation; the influence of assembled structures on expression is similarly indicated. Assembly begins with the production and then activation, via phosphorylation, of CtrA in response to (as-yet-unknown) cell cycle cues. The expression of class III genes is dependent upon class II gene products and assembly of the early basal-body structures. Mutations at the bfa locus bypass this requirement. It is hypothesized that bfa regulation of class III genes occurs, because the Bfa protein is exported from the cell solely by the assembled early basal-body structures and can no longer be exported once the class III structures are added. Thus, during this time Bfa concentrations would be low in the cell, allowing class III gene transcription. Before and after this window, high levels of the Bfa protein would serve to repress class III genes. Similarly, synthesis of the class IV flagellins is coupled to successful assembly of class III structures, although in this case the regulatory mechanism is known to be posttranscriptional. Apparently, FlbT destabilizes the flagellin mRNA, either directly or indirectly, thus significantly reducing what is otherwise an extremely long half-life. Completion of the basal body and hook assembly seems to counteract FlbT activity and allows translation of the flagellin mRNA.

Comparison of known cis-acting regulatory elements of class II and class III genes. Shown are the intergenic regions between the fliX gene and the flgI operon and between the fliL and flgF operons, as well as the upstream regulatory region of the fliF operon. The positions of the promoters (σ70 for class II and σ54 for class III) and cis-acting sequences are also diagrammed, with the positive or negative regulation of the corresponding binding factor indicated by an arrow or barred line, respectively. FlbD has been shown to down-regulate the class II fliF operon, perhaps by competing with the activator CtrA. FlbD activates the class III operons, presumably brought into proximity with the promoters by an IHF-induced bending of the intervening DNA. It has been speculated that the close, or partially overlapping, organization of ftr and CtrA box elements in the intergenic regions provides a means of coordinating the expression of these class II and class III genes.

Polarity is established in the C. crescentus predivisional cell as a result of compartmentalized gene expression or protein targeting. The schematic diagrams of a portion of the C. crescentus cell cycle show the early and late predivisional cells and the resulting progeny cells. (A) Relative expression of flagellar genes in the two poles and progeny cells. Transcription of the late flagellar genes (classes III and IV) exclusively in the swarmer pole of the predivisional cell is due to the swarmer pole-specific activation of the transcriptional activator, FlbD. Following the biogenesis of the cell division plane, FlbD also negatively regulates the class II fliF promoter in the swarmer pole, which is therefore only transcribed in the stalked pole, where FlbD is inactive. Because late class IV flagellin genes are only transcribed in the swarmer pole of the predivisional cell after the cell division plane is intact, the long-lived flagellin mRNA is segregated to the swarmer progeny cell, ensuring that the still-growing flagellar filament will have adequate flagellin precursors. FlbE is also necessary for the correct pole-specific program of flagellar gene expression and has been found to localize to the stalked pole and to the cell division plane in the swarmer compartment. The protein, with regions of homology to known sensor histidine kinases, is able to transfer phosphate groups to FlbD in vitro. This evidence, and the correlation between FlbE localization and FlbD activation in the swarmer pole, suggests the attractive possibility that FlbE, triggered by its localization to structures at the cell division plane, maintains FlbD activation by phosphorylation. FlbE localized to the stalked pole is apparently unable to phosphorylate FlbD. (B) Establishment of MCP localization. MCP is synthesized in the predivisional cell and targeted to the swarmer pole. The MCP in the swarmer cell is later degraded at the swarmer-to-stalked-cell transition. Regions of the carboxyl terminus of the protein direct both polar localization and proteolytic degradation.