Flagellar Gene Expression Research Articles

CsrA impacts survival of Yersinia enterocolitica by affecting a myriad of physiological activities.

BackgroundA previous study identified a Yersinia enterocolitica transposon mutant, GY448, that was unable to export the flagellar type three secretion system (T3SS)-dependent phospholipase, YplA. This strain was also deficient for motility and unable to form colonies on Lauria-Bertani agar medium. Preliminary analysis suggested it carried a mutation in csrA. CsrA in Escherichia coli is an RNA-binding protein that is involved in specific post-transcriptional regulation of a myriad of physiological activities. This study investigated how CsrA affects expression of the flagellar regulatory cascade that controls YplA export and motility. It also explored the effect of csrA mutation on Y. enterocolitica in response to conditions that cue physiological changes important for growth in environments found both in nature and the laboratory.ResultsThe precise location of the transposon insertion in GMY448 was mapped within csrA. Genetic complementation restored disruptions in motility and the YplA export phenotype (Yex), which confirmed this mutation disrupted CsrA function. Mutation of csrA affected expression of yplA and flagellar genes involved in flagellar T3SS dependent export and motility by altering expression of the master regulators flhDC. Mutation of csrA also resulted in increased sensitivity of Y. enterocolitica to various osmolytes, temperatures and antibiotics.ConclusionsThe results of this study reveal unique aspects of how CsrA functions in Y. enterocolitica to control its physiology. This provides perspective on how the Csr system is susceptible to adaptation to particular environments and bacterial lifestyles.

Mathematical model of flagella gene expression dynamics in Salmonella enterica serovar typhimurium

Flagellar assembly in Salmonella is controlled by an intricate genetic and biochemical network. This network comprises of a number of inter-connected feedback loops, which control the assembly process dynamically. Critical among these are the FliA-FlgM feedback, FliZ-mediated positive feedback, and FliT-mediated negative feedback. In this work, we develop a mathematical model to track the dynamics of flagellar gene expression in Salmonella. Analysis of our model demonstrates that the network is wired to not only control the transition of the cell from a non-flagellated to a flagellated state, but to also control dynamics of gene expression during cell division. Further, we predict that FliZ encoded in the flagellar regulon acts as a critical secretion-dependent molecular link between flagella and Salmonella Pathogenicity Island 1 gene expression. Sensitivity analysis of the model demonstrates that the flagellar regulatory network architecture is extremely robust to mutations.

Role of VcrD1 protein in expression and secretion of flagellar components in Vibrio parahaemolyticus.

VcrD1 protein is a component of type III secretion system (T3SS) 1 in Vibrio parahaemolyticus. A comparative analysis of secretomes of wild-type and ΔvcrD1 strains revealed that the mutant was defective in secretion of diverse proteins including several flagellar components. Western blot analyses using specific antibodies confirmed that the secretion of at least four flagellar components, such as FlaA, FlgL, FlgE, and FlgM, was affected by the vcrD1 mutation, which was consistent with decreased motility on soft agar plates and the non-flagellated morphology of the mutant. The ΔexsA mutant, another T3SS1 mutant, did not showed reduced motility, but became non-motile phenotype with the additional ΔvcrD1 mutation. Complementation of wild-type vcrD1 gene into ΔvcrD1 mutant resulted in restored motility. Fractionation of bacterial cytoplasm from the periplasm and membrane revealed lower levels of FlaA and FlgM in the cytoplasm of the ΔvcrD1 mutant, indicating that VcrD1 might regulate the expression of flagellar genes in addition to the secretion of flagellar components in V. parahaemolyticus. Quantitative RT-PCR assays of seven representative flagellar genes in the wild-type and ΔvcrD1 mutant strains demonstrated that transcript levels of two early flagellar genes, flaK and flaL, were not reduced by the vcrD1 mutation, whereas the middle and late flagellar genes were expressed at a lower level in the vcrD1 mutant. This study raises a possibility that VcrD1 plays a role in flagellar morphogenesis in V. parahaemolyticus by regulating the expression and secretion of flagellar components.

Induction of the lateral flagellar system of Vibrio shilonii is an early event after inhibition of the sodium ion flux in the polar flagellum.

In this study, we show the induction of lateral flagella by the action of the sodium channel blocker phenamil, in the marine bacterium Vibrio shilonii, a coral pathogen that causes bleaching. We analyzed the growth and morphology of cells treated with phenamil. A time course analysis showed that after 30 min of exposure to the sodium channel blocker, lateral flagella were present and could be detected by electron microscopy. Detection of the mRNA of the master regulator (lafK) and lateral flagellin (lafA) by RT-PCR confirmed the expression of lateral flagellar genes. We show the simultaneous isolation of polar and, for the first time, lateral flagellar hook-basal bodies. This allowed us to compare the dimensions and morphological characteristics of the 2 structures.

Loss of the lac operon contributes to Salmonella invasion of epithelial cells through derepression of flagellar synthesis.

Salmonella, a genus that is closely related to Escherichia coli, includes many pathogens of humans and other animals. A notable feature that distinguishes Salmonella from E. coli is lactose negativity, because the lac operon is lost in most Salmonella genomes. Here, we expressed the lac operon in Salmonella enterica serovar Typhimurium and compared the virulence of the Lac(+) strain to that of the wild-type strain in a murine model, invasion assays, and macrophage replication assays. We showed that the Lac(+) strain is attenuated in vivo and the attenuation of virulence is caused by its defect in epithelial cell invasion. However, the invasion-defective phenotype is unrelated to lactose utilization. Through sequencing and the comparison of the transcriptome profile between the Lac(+) and wild-type strains during invasion, we found that most flagellar genes were markedly downregulated in the Lac(+) strain, while other genes associated with invasion, such as the majority of genes encoded in Salmonella pathogenicity island 1, were not differentially expressed. Moreover, we discovered that lacA is the major repressor of flagellar gene expression in the lac operon. In conclusion, these data demonstrate that the lac operon decreases Salmonella invasion of epithelial cells through repression of flagellar biosynthesis. As the ability to invade epithelial cells is a critical virulence determinant of Salmonella, our results provide important evidence that the loss of the lac operon contributes to the evolution of Salmonella pathogenicity.

FliT Selectively Enhances Proteolysis of FlhC Subunit in FlhD4C2 Complex by an ATP-dependent Protease, ClpXP

We previously reported that the ClpXP ATP-dependent protease specifically recognizes and degrades the flagellar master transcriptional activator complex, FlhD4C2, to negatively control flagellar biogenesis. The flagellum-related protein, FliT, is also a negative regulator of flagellar regulon by inhibiting the binding of FlhD4C2 to the promoter DNA. We have found a novel pathway of FliT inhibition of FlhD4C2 activity connected to ClpXP proteolysis. An in vitro degradation assay using purified proteins shows that FliT selectively increases ClpXP proteolysis of the FlhC subunit in the FlhD4C2 complex. FliT behaves specifically to ClpXP-dependent proteolysis of FlhC. An in vitro interaction assay detects the ternary complex of FliT-FlhD4C2-ClpX. FliT promotes the affinity of ClpX against FlhD4C2 complex, whereas FliT does not directly interact with ClpX. Thus, FliT interacts with the FlhC in FlhD4C2 complex and increases the presentation of the FlhC recognition region to ClpX. The DNA-bound form of FlhD4C2 complex is resistant to ClpXP proteolysis. We suggest that the role of FliT in negatively controlling the flagellar gene expression involves increasing free molecules of FlhD4C2 sensitive to ClpXP proteolysis by inhibiting the binding to the promoter DNA as well as enhancing the selective proteolysis of FlhC subunit by ClpXP.

Mutually repressing repressor functions and multi-layered cellular heterogeneity regulate the bistable Salmonella fliC census.

Bistable flagellar and virulence gene expression generates specialized Salmonella subpopulations with distinct functions. Repressing flagellar genes allows Salmonella to evade caspase-1 mediated host defenses and enhances systemic colonization. By definition, bistability arises when intermediate states of gene expression are rendered unstable by the underlying genetic circuitry. We demonstrate sustained bistable fliC expression in virulent Salmonella 14028 and document dynamic control of the distribution, or single-cell census, of flagellar gene expression by the mutually repressing repressors YdiV and FliZ. YdiV partitions cells into the fliC-OFF subpopulation, while FliZ partitions cells into the fliC-HIGH subpopulation at late time points during growth. Bistability of ΔfliZ populations and ydiV-independent FliZ control of flagellar gene expression provide evidence that the YdiV-FliZ mutually repressing repressor circuit is not required for bistability. Repression and activation by YdiV and FliZ (respectively) can shape the census of fliC expression independently, and bistability collapses into a predominantly intermediate population in the absence of both regulators. Metered expression of YdiV and FliZ reveals variable sensitivity to these regulators and defines conditions where expression of FliZ enhances fliC expression and where FliZ does not alter the fliC census. Thus, this evolved genetic circuitry coordinates multiple layers of regulatory heterogeneity into a binary response.

FlgM is secreted by the flagellar export apparatus in Bacillus subtilis.

The bacterial flagellum is assembled from over 20 structural components, and flagellar gene regulation is morphogenetically coupled to the assembly state by control of the anti-sigma factor FlgM. In the Gram-negative bacterium Salmonella enterica, FlgM inhibits late-class flagellar gene expression until the hook-basal body structural intermediate is completed and FlgM is inhibited by secretion from the cytoplasm. Here we demonstrate that FlgM is also secreted in the Gram-positive bacterium Bacillus subtilis and is degraded extracellularly by the proteases Epr and WprA. We further demonstrate that, like in S. enterica, the structural genes required for the flagellar hook-basal body are required for robust activation of σ(D)-dependent gene expression and efficient secretion of FlgM. Finally, we determine that FlgM secretion is strongly enhanced by, but does not strictly require, hook-basal body completion and instead demands a minimal subset of flagellar proteins that includes the FliF/FliG basal body proteins, the flagellar type III export apparatus components FliO, FliP, FliQ, FliR, FlhA, and FlhB, and the substrate specificity switch regulator FliK.

Alternative sigma factor RpoN2 is required for flagellar motility and full virulence of Xanthomonas oryzae pv. oryzae

Xanthomonas oryzae pv. oryzae (Xoo), the bacterial blight pathogen of rice, harbors a single polar flagellum for motility. How the flagellar system is regulated and how it is related to bacterial pathogenesis are not well understood. The genomic sequence of Xoo strain PXO99A revealed a flagellar regulon containing over 60 contiguous genes. A gene encoding alternative sigma factor 54 (σ54), named as rpoN2, is located in the central region of the regulon. RT-PCR analysis demonstrated rpoN2 was transcribed in an operon with flgRR, and fleQ. Single gene deletion mutants of the rpoN2 operon were generated. The rpoN2 and fleQ mutant lost swimming motility, whereas the flgRR mutant remained motile. Quantitative RT-PCR analysis further demonstrated that expression of regulatory genes fliA and flgM, and structural genes flgG, flhB, and fliC were significantly down-regulated in the rpoN2 and fleQ mutants. These results indicated that RpoN2 and FleQ synergistically controlled flagellar motility by regulating gene expression. Interestingly, the rpoN2 mutant, but not the fleQ mutant was impaired in its virulence on rice. In addition, we showed that the flagellin gene fliC mutant, which was non-motile, was not defective in virulence. Thus, we concluded that flagellar motility might not be essential for Xoo virulence in rice, and RpoN2 probably regulated bacterial virulence through a manner independent of its role in controlling flagellar gene expression.

Requirement of the flagellar protein export apparatus component FliO for optimal expression of flagellar genes in Helicobacter pylori.

Flagellar biogenesis in Helicobacter pylori involves the coordinated expression of flagellar genes with assembly of the flagellum. The H. pylori flagellar genes are organized into three regulons based on the sigma factor needed for their transcription (RpoD [σ(80)], RpoN [σ(54)], or FliA [σ(28)]). Transcription of RpoN-dependent genes is activated by a two-component system consisting of the sensor kinase FlgS and the response regulator FlgR. While the cellular cues sensed by the FlgS/FlgR two-component system remain to be elucidated, previous studies revealed that disrupting certain components of the flagellar export apparatus inhibited transcription of the RpoN regulon. FliO is the least conserved of the membrane-bound components of the export apparatus and has not been annotated for any of the H. pylori genomes sequenced to date. A PSI-BLAST analysis identified a potential H. pylori FliO protein which membrane topology algorithms predict to possess a large N-terminal periplasmic domain that is absent from FliO of Escherichia coli and Salmonella, the paradigms for flagellar structure/function studies. FliO was necessary for flagellar biogenesis as well as wild-type levels of motility and transcription of RpoN-dependent and FliA-dependent flagellar genes in H. pylori strain B128. FliO also appears to be required for wild-type levels of the export apparatus protein FlhA in the membrane. Interestingly, the periplasmic and cytoplasmic domains were somewhat dispensable for flagellar gene regulation and assembly, suggesting that these domains have relatively minor roles in flagellar synthesis.

A conservative amino acid mutation in the master regulator FleQ renders Pseudomonas aeruginosa aflagellate.

Flagellar-based motility plays a critical role in Pseudomonas aeruginosa pathogenesis, influencing both the establishment of bacterial infection and the host's response to the pathogen. Nonetheless, aflagellate clinical strains are often isolated from acutely and chronically infected patients and include the virulent laboratory strain PA103. We determined that PA103's aflagellate phenotype is the result of a single amino acid change (G240V) in the master flagellar regulator, FleQ. This mutation, which lies just outside the Walker B box of FleQ, abrogates the ability of FleQ to positively regulate flagellar gene expression. Reversal of this seemingly conservative amino acid substitution is sufficient to restore swimming motility to PA103, despite the presence of mutations in other flagellar genes of PA103. We also investigated the consequences of restoring flagellar assembly on PA103 virulence. Although a negative correlation between flagellar assembly and Type 3 secretion system (T3SS) expression has been reported previously, we did not observe downregulation of T3SS expression or function in Fla+ PA103. Restoration of flagellar assembly did, however, amplify IL-1 signals measured during murine pulmonary infection and was associated with increased bacterial clearance. These experiments suggest that loss of flagellar motility may primarily benefit PA103 by attenuating pathogen recognition and clearance during acute infection.

Analysis of Factors That Affect FlgM-Dependent Type III Secretion for Protein Purification with Salmonella enterica Serovar Typhimurium

The FlgM protein is secreted in response to flagellar hook-basal body secretion and can be used as a secretion signal to direct selected protein secretion via the flagellar type III secretion (T3S) system [H. M. Singer, M. Erhardt, A. M. Steiner, M. M. Zhang, D. Yoshikami, G. Bulaj, B. M. Olivera, and K. T. Hughes, mBio 3(3):e00115-12, 2012, http://dx.doi.org/10.1128/mBio.00115-12]. Conditions known to affect flagellar gene expression, FlgM stability, and flagellar T3S were tested either alone or in combination to determine their effects on levels of secreted FlgM. These conditions included mutations that affect activity of the flagellar FlhD4C2 master regulatory protein complex or the FlgM T3S chaperone σ(28), the removal of Salmonella pathogenicity island 1 (Spi1), the removal of flagellar late secretion substrates that could compete with FlgM for secretion, and changes in the ionic strength of the growth medium. Conditions that enhanced FlgM secretion were combined in order to maximize levels of secreted FlgM. An optimized FlgM secretion strain was used to secrete and isolate otherwise difficult-to-produce proteins and peptides fused to the C terminus of FlgM. These include cysteine-rich, hydrophobic peptides (conotoxins δ-SVIE and MrVIA), nodule-specific, cysteine-rich antimicrobial peptides (NCR), and a malaria surface antigen domain of apical membrane antigen AMA-1.

Molecular Characterization of “Inconsistent” Variants ofSalmonellaTyphimurium Isolated in Italy

Salmonella 4,[5],12:i:- is a variant of Salmonella Typhimurium, which lacks the expression of phase-2 flagellar antigen, generally associated with the deletion of the fljB gene. Additional mechanisms involving the fljAB operon ( fljA, fljB, and hin genes) lead to the lack of expression of phase-2 flagellar antigens also in Salmonella strains harboring the fljB gene. For 20 S. 4,[5],12:i:- strains, defined as "inconsistent" Salmonella Typhimurium variants since they had phenotypically behaved as monophasic, even though the fljB gene was conserved, the fljAB operon was characterized in order to explain the ineffective expression of the phase-2 flagellar antigen. The monophasic phenotype for a first group of strains (9) was likely due to the absence of the hin gene, leading to the inhibited switch between the expression of phase-1 and phase-2 flagellar genes. For a second group of strains (5), the monophasic phenotype could be attributed to nonconservative point mutations identified in fljA and hin genes, which could hamper the proper expression of invertase gene and the fljA, acting as repressor of the phase-1 flagellar gene. Finally, for a last group of inconsistent strains (6), a plausible reason for their monophasic phenotype was not found, since the genes involved in the expression of phase-2 flagellar antigen were fully conserved. Moreover, the collection of inconsistent Salmonella Typhimurium isolates investigated were characterized by distinct molecular profiles, as demonstrated by multiple-locus variable-number tandem repeat analysis, and phenotype variability, as demonstrated by phage-typing. This study highlights the usefulness of investigating the entire fljAB operon when a definitive identification of the monophasic or biphasic status of Salmonella Typhimurium strains is needed (for instance, in the context of epidemiological investigations aimed to identify the relatedness among strains).

Quorum sensing positively regulates flagellar motility in pathogenic Vibrio harveyi

Vibrios belonging to the Harveyi clade are among the major pathogens of aquatic organisms. Quorum sensing (QS) is essential for virulence of V. harveyi towards different hosts. However, most virulence factors reported to be controlled by QS to date are negatively regulated by QS, therefore suggesting that their impact on virulence is limited. In this study, we report that QS positively regulates flagellar motility. We found that autoinducer synthase mutants showed significantly lower swimming motility than the wild type, and the swimming motility could be restored by adding synthetic signal molecules. Further, motility of a luxO mutant with inactive QS (LuxO D47E) was significantly lower than that of the wild type and of a luxO mutant with constitutively maximal QS activity (LuxO D47A). Furthermore, we found that the expression of flagellar genes (both early, middle and late genes) was significantly lower in the luxO mutant with inactive QS when compared with wild type and the luxO mutant with maximal QS activity. Motility assays and gene expression also revealed the involvement of the quorum-sensing master regulator LuxR in the QS regulation of motility. Finally, the motility inhibitor phenamil significantly decreased the virulence of V. harveyi towards gnotobiotic brine shrimp larvae.

The effect of cell growth phase on the regulatory cross-talk between flagellar and Spi1 virulence gene expression.

The flagellar regulon controls Salmonella biofilm formation, virulence gene expression and the production of the major surface antigen present on the cell surface: flagellin. At the top of a flagellar regulatory hierarchy is the master operon, flhDC, which encodes the FlhD4C2 transcriptional complex required for the expression of flagellar, chemotaxis and Salmonella pathogenicity island 1 (Spi1) genes. Of six potential transcriptional start-sites within the flhDC promoter region, only two, P1flhDC and P5flhDC, were functional in a wild-type background, while P6flhDC was functional in the absence of CRP. These promoters are transcribed differentially to control either flagellar or Spi1 virulent gene expression at different stages of cell growth. Transcription from P1flhDC initiates flagellar assembly and a negative autoregulatory loop through FlhD4C2-dependent transcription of the rflM gene, which encodes a repressor of flhDC transcription. Transcription from P1flhDC also initiates transcription of the Spi1 regulatory gene, hilD, whose product, in addition to activating Spi1 genes, also activates transcription of the flhDC P5 promoter later in the cell growth phase. The regulators of flhDC transcription (RcsB, LrhA, RflM, HilD, SlyA and RtsB) also exert their control at different stages of the cell growth phase and are also subjected to cell growth phase control. This dynamic of flhDC transcription separates the roles of FlhD4C2 transcriptional activation into an early cell growth phase role for flagellar production from a late cell growth phase role in virulence gene expression.

Crystallization of the N-terminal regulatory domain of the enhancer-binding protein FleQ fromStenotrophomonas maltophilia

FleQ is a master regulator that controls bacterial flagellar gene expression. It is a unique enhancer-binding protein or repressor protein comprising an N-terminal FleQ domain, an AAA(+)/ATPase σ54-interaction domain and a helix-turn-helix DNA-binding domain. FleN is a putative ATPase with a deviant Walker A motif that works together with FleQ by binding to the FleQ N-terminal domain to fully express pel, psl and cdr operons in the presence of c-di-GMP to enhance biofilm formation. Stenotrophomonas maltophilia is an emerging human pathogen that causes fatal infections in humans. In order to understand the interaction between the FleN and FleQ domains and its effect on S. maltophilia biofilm formation, determination of the FleQ-c-di-GMP and FleN-FleQ-c-di-GMP complex structures was embarked upon. Towards this goal, the FleQ N-terminal domain from S. maltophilia was first cloned and expressed in Escherichia coli. Native and SeMet-labelled FleQ domains were successfully crystallized and diffracted to resolutions of 2.08 and 2.58 Å, respectively.

The Salmonella Spi1 Virulence Regulatory Protein HilD Directly Activates Transcription of the Flagellar Master OperonflhDC

Infection of intestinal epithelial cells is dependent on the Salmonella enterica serovar Typhimurium pathogenicity island 1 (Spi1)-encoded type III injectisome system and flagellar motility. Thus, the expression of virulence and flagellar genes is subject to tight regulatory control mechanisms in order to ensure the correct spatiotemporal production of the respective gene products. In this work, we reveal a new level of cross-regulation between the Spi1 and flagellar regulatory systems. Transposon mutagenesis identified a class of mutants that prevented flhDC autorepression by overexpressing HilD. HilD, HilC, RtsA, and HilA comprise a positive regulatory circuit for the expression of the Spi1 genes. Here, we report a novel transcriptional cross talk between the Spi1 and flagellar regulons where HilD transcriptionally activates flhDC gene expression by binding to nucleotides -68 to -24 upstream from the P5 transcriptional start site. We additionally show that, in contrast to the results of a previous report, HilA does not affect flagellar gene expression. Finally, we discuss a model of the cross-regulation network between Spi1 and the flagellar system and propose a regulatory mechanism via the Spi1 master regulator HilD that would prime flagellar genes for rapid reactivation during host infection.

Quorum Sensing Controls Flagellar Morphogenesis in Burkholderia glumae

Burkholderia glumae is a motile plant pathogenic bacterium that has multiple polar flagella and one LuxR/LuxI-type quorum sensing (QS) system, TofR/TofI. A QS-dependent transcriptional regulator, QsmR, activates flagellar master regulator flhDC genes. FlhDC subsequently activates flagellar gene expression in B. glumae at 37°C. Here, we confirm that the interplay between QS and temperature is critical for normal polar flagellar morphogenesis in B. glumae. In the wild-type bacterium, flagellar gene expression and flagellar number were greater at 28°C compared to 37°C. The QS-dependent flhC gene was significantly expressed at 28°C in two QS-defective (tofI::Ω and qsmR::Ω) mutants. Thus, flagella were present in both tofI::Ω and qsmR::Ω mutants at 28°C, but were absent at 37°C. Most tofI::Ω and qsmR::Ω mutant cells possessed polar or nonpolar flagella at 28°C. Nonpolarly flagellated cells processing flagella around cell surface of both tofI::Ω and qsmR::Ω mutants exhibited tumbling and spinning movements. The flhF gene encoding GTPase involved in regulating the correct placement of flagella in other bacteria was expressed in QS mutants in a FlhDC-dependent manner at 28°C. However, FlhF was mislocalized in QS mutants, and was associated with nonpolar flagellar formation in QS mutants at 28°C. These results indicate that QS-independent expression of flagellar genes at 28°C allows flagellar biogenesis, but is not sufficient for normal polar flagellar morphogenesis in B. glumae. Our findings demonstrate that QS functions together with temperature to control flagellar morphogenesis in B. glumae.

Post-Transcriptional and Post-Translational Control of the Flagellar Regulon Rescues Motility of a Salmonella Enterica Type III Export Flio Mutant

The bacterial flagellum is made using a type III secretion pathway. For Salmonella enterica, this requires 6 transmembrane proteins FlhA, FlhB, FliO, FliP, FliQ, and FliR. FliO is not absolutely required, because bypass mutations in the fliP gene can improve motility for a fliO mutant. In this study, an extended screen was made for randomly selected mutations that improved motility of cells bearing a ΔfliO null mutation. Using whole genome sequencing two novel mutations were identified, a “silent” mutation localized to the fliA gene, which encoded a U36C substitution in the fliAZ mRNA. A missense mutation was found in the clpP gene, and encoded a V20F substitution in the ClpXP protease. Transcriptional and translational fusions of the lacZ gene to the fliA promoter demonstrated that the silent mutation improved translation of the highly structured fliAZ mRNA. Real-time quantitative RT-PCR revealed that late flagellar gene expression was blocked in a ΔfliO mutant, but the clpP(V20F) or fliA(mRNA U36C) mutations improved expression. Examination of flagellar biogenesis by immunoblotting and transmission electron microscopy found that the fliO mutant synthesized rare flagella, but introducing the fliA(mRNA U36C) mutation together with previously identified mutations in fliP cooperatively restored flagellar biogenesis to near wild-type levels.

Themes and Variations: Regulation of RpoN-Dependent Flagellar Genes across Diverse Bacterial Species

Flagellar biogenesis in bacteria is a complex process in which the transcription of dozens of structural and regulatory genes is coordinated with the assembly of the flagellum. Although the overall process of flagellar biogenesis is conserved among bacteria, the mechanisms used to regulate flagellar gene expression vary greatly among different bacterial species. Many bacteria use the alternative sigma factor σ 54 (also known as RpoN) to transcribe specific sets of flagellar genes. These bacteria include members of the Epsilonproteobacteria (e.g., Helicobacter pylori and Campylobacter jejuni), Gammaproteobacteria (e.g., Vibrio and Pseudomonas species), and Alphaproteobacteria (e.g., Caulobacter crescentus). This review characterizes the flagellar transcriptional hierarchies in these bacteria and examines what is known about how flagellar gene regulation is linked with other processes including growth phase, quorum sensing, and host colonization.