# Bacterial Flagellar Filament: A Supramolecular Multifunctional Nanostructure
## 5 Protein Synthesis, Export and Filament Elongation  
### Genetic Regulation of FliC and FliD  
The genetic organization of the flagellar components is highly complex and relatively well conserved across bacteria with the same type of flagellar arrangement (i.e., peritrichuous, monotrichous/polar, etc.) but quite different between them [1,99,100]. The bacterial flagellar regulon commonly consists of dozens of genes grouped in several persons, encoding all structural proteins of the flagellum, the chemosensory apparatus and the regulators that control gene expression. For example, there are about 50 genes involved in the synthesis of the polar _P. aeruginosa_ flagella, distributed in 17 putative operons comprising 41 flagellar genes (26 encoding structural proteins, eight encoding regulators and six involved in the export apparatus), linked to nine genes involved in chemotaxis and clustered in three regions of the chromosome constituting the _fla_ regulon [101,102]. On the other hand, in peritrichous bacteria such as _Salmonella_ spp. and _Escherichia coli_, the flagellar regulon contains 70 genes distributed in at least 25 operons [79,99]. The transcription of these genes is hierarchical and controlled by different promoter classes that are temporally regulated (Figure 4, Table 1). In peritrichous bacteria, flagellar genes are organized into three classes (I-III), starting with the transcription of the class I master operon as a response to the environmental stimuli that leads to the synthesis of the transcriptional regulator FlhDC. This regulator, in turn, activates the transcription of the class II genes responsible for the assembly of the hook-basal body (HBB) complex. Among class II genes are two crucial regulators of the class III genes: transcription factor FliA (\(\alpha^{28}\)) responsible for the transcription of class III genes, and FlgM, which acts as a FliA inhibitor. Once the HBB is assembled, FlgM is exported out of the cell via the HBB central channel, releasing FliA to interact with RNA polymerase and guide the transcription of the class III genes, including flagellin. This three-tiered organization ensures that cells produce flagella in response to the environmental signals and that energetically costly synthesis of the filament  
Figure 3: FliD and the filament cap. (**a**) Structure of FliD from _E. coli_ shows three domains (D1–D3) (PDB: 5H5V). The N- and C-termini are highly flexible and could not be resolved. (**b**) Structural alignment of FliD molecules from different species show high structural homology of D2 and D3 domains. FliD from _H. pylori_ has two additional domains, D4 and D5. (**c**) Crystal structure of the filament cap from _E. coli_ showing a hexameric structure. Domains D2 and D3 form the head region, while D1 (incomplete) forms the flexible leg region; (**d**) Oligomeric states of the crystallized FliD caps differ across species. proceeds only when enough HBBs have been formed. An exception is the _Actinoplanes missouriensis_ zoospore in which there is no hierarchical coordination and 33 flagellar genes are transcribed simultaneously during the sporangium formation [103]. The filament cap gene _filD_ belongs to the group of class II genes, whereas flagellin genes belong to class III. Although _filD_ is expressed as a class II gene, it is not assembled into the growing flagellar structure until the hook and the junction proteins are expressed from class III (including flagellin genes) and incorporated into the flagellar organelle [104]. Hence, this cascade serves to control the timing of gene expression to coincide with the assembly of the flagellar apparatus and filament. As mentioned above, \(S\). Typhimurium exhibits phase variation, or alternate expression of two flagellin genes, _filC_ and _filB_. This process is regulated by a control region upstream of _filB_ that is a subject of inversion, which orients the region forward and generates a promoter. This allows co-transcription of _filB_ and _filA_, a transcriptional repressor that inhibits the expression of FliC [105].