scholarly journals Coupling of Flagellar Gene Expression to Flagellar Assembly in Salmonella enterica Serovar Typhimurium andEscherichia coli

2000 ◽  
Vol 64 (4) ◽  
pp. 694-708 ◽  
Author(s):  
Gavin S. Chilcott ◽  
Kelly T. Hughes
Keyword(s):  
Basal Body ◽  
Cell Cycle Control ◽  
Regulatory Protein ◽  
Intermediate Structure ◽  
Flagellar Gene ◽  
Transcriptional Regulatory Mechanism ◽  
Transcriptional Regulatory ◽  
Serovar Typhimurium ◽  
Ring Structures ◽  
Flagellar Assembly

SUMMARY How do organisms assess the degree of completion of a large structure, especially an extracellular structure such as a flagellum? Bacteria can do this. Mutants that lack key components needed early in assembly fail to express proteins that would normally be added at later assembly stages. In some cases, the regulatory circuitry is able to sense completion of structures beyond the cell surface, such as completion of the external hook structure. In Salmonella and Escherichia coli, regulation occurs at both transcriptional and posttranscriptional levels. One transcriptional regulatory mechanism involves a regulatory protein, FlgM, that escapes from the cell (and thus can no longer act) through a complete flagellum and is held inside when the structure has not reached a later stage of completion. FlgM prevents late flagellar gene transcription by binding the flagellum-specific transcription factor ς28. FlgM is itself regulated in response to the assembly of an incomplete flagellum known as the hook-basal body intermediate structure. Upon completion of the hook-basal body structure, FlgM is exported through this structure out of the cell. Inhibition of ς28-dependent transcription is relieved, and genes required for the later assembly stages are expressed, allowing completion of the flagellar organelle. Distinct posttranscriptional regulatory mechanisms occur in response to assembly of the flagellar type III secretion apparatus and of ring structures in the peptidoglycan and lipopolysaccharide layers. The entire flagellar regulatory pathway is regulated in response to environmental cues. Cell cycle control and flagellar development are codependent. We discuss how all these levels of regulation ensure efficient assembly of the flagellum in response to environmental stimuli.

scholarly journals FliZ Is a Posttranslational Activator of FlhD4C2-Dependent Flagellar Gene Expression

2008 ◽  
Vol 190 (14) ◽  
pp. 4979-4988 ◽  
Author(s):  
Supreet Saini ◽  
Jonathon D. Brown ◽  
Phillip D. Aldridge ◽  
Christopher V. Rao
Keyword(s):  
Gene Expression ◽  
Sigma Factor ◽  
Flagellar Gene ◽  
Critical Aspect ◽  
Swarming Motility ◽  
Flagellar Biosynthesis ◽  
Serovar Typhimurium ◽  
Flagellar Assembly

ABSTRACT 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 σ28-FlgM checkpoint, which couples the expression of late flagellar (Pclass3) genes to the completion of the hook-basal body. In this work, we investigated the role of FliZ in coupling middle flagellar (Pclass2) gene expression to assembly in Salmonella enterica serovar Typhimurium. We demonstrate that FliZ is an FlhD4C2-dependent activator of Pclass2/middle gene expression. Our results suggest that FliZ regulates the concentration of FlhD4C2 posttranslationally. We also demonstrate that FliZ functions independently of the flagellum-specific sigma factor σ28 and the filament-cap chaperone/FlhD4C2 inhibitor FliT. Furthermore, we show that the previously described ability of σ28 to activate Pclass2/middle gene expression is, in fact, due to FliZ, as both are expressed from the same overlapping Pclass2 and Pclass3 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.

scholarly journals Oligomerization of the FliF Domains Suggests a Coordinated Assembly of the Bacterial Flagellum MS Ring

2022 ◽  
Vol 12 ◽  
Author(s):  
Giuseppina Mariano ◽  
Raquel Faba-Rodriguez ◽  
Soi Bui ◽  
Weilong Zhao ◽  
James Ross ◽  
...  
Keyword(s):  
Basal Body ◽  
Computational Analysis ◽  
Cell Envelope ◽  
Bacterial Flagellum ◽  
Self Assembling ◽  
Single Protein ◽  
Serovar Typhimurium ◽  
Ring Structures ◽  
Periplasmic Domain ◽  
The Individual

The bacterial flagellum is a complex, self-assembling macromolecular machine that powers bacterial motility. It plays diverse roles in bacterial virulence, including aiding in colonization and dissemination during infection. The flagellum consists of a filamentous structure protruding from the cell, and of the basal body, a large assembly that spans the cell envelope. The basal body is comprised of over 20 different proteins forming several concentric ring structures, termed the M- S- L- P- and C-rings, respectively. In particular, the MS rings are formed by a single protein FliF, which consists of two trans-membrane helices anchoring it to the inner membrane and surrounding a large periplasmic domain. Assembly of the MS ring, through oligomerization of FliF, is one of the first steps of basal body assembly. Previous computational analysis had shown that the periplasmic region of FliF consists of three structurally similar domains, termed Ring-Building Motif (RBM)1, RBM2, and RBM3. The structure of the MS-ring has been reported recently, and unexpectedly shown that these three domains adopt different symmetries, with RBM3 having a 34-mer stoichiometry, while RBM2 adopts two distinct positions in the complex, including a 23-mer ring. This observation raises some important question on the assembly of the MS ring, and the formation of this symmetry mismatch within a single protein. In this study, we analyze the oligomerization of the individual RBM domains in isolation, in the Salmonella enterica serovar Typhimurium FliF ortholog. We demonstrate that the periplasmic domain of FliF assembles into the MS ring, in the absence of the trans-membrane helices. We also report that the RBM2 and RBM3 domains oligomerize into ring structures, but not RBM1. Intriguingly, we observe that a construct encompassing RBM1 and RBM2 is monomeric, suggesting that RBM1 interacts with RBM2, and inhibits its oligomerization. However, this inhibition is lifted by the addition of RBM3. Collectively, this data suggest a mechanism for the controlled assembly of the MS ring.

scholarly journals Negative transcriptional regulation in the Caulobacter flagellar hierarchy

1989 ◽  
Vol 86 (17) ◽  
pp. 6656-6660 ◽  
Author(s):  
H Xu ◽  
A Dingwall ◽  
L Shapiro
Keyword(s):  
Gene Expression ◽  
Basal Body ◽  
Caulobacter Crescentus ◽  
Negative Control ◽  
Negative Regulation ◽  
Flagellar Gene ◽  
Structural Genes ◽  
Number Of Genes ◽  
Flagellar Assembly ◽  
Transcriptional Fusions

The Caulobacter crescentus flagellum is formed at a specific time in the cell cycle and its assembly requires the ordered expression of a large number of genes. These genes are controlled in a positive trans-acting hierarchy that reflects the order of assembly of the flagellum. Using plasmids carrying transcriptional fusions of either a neo or a lux reporter gene to the promoters of three flagellar genes representing different ranks in the hierarchy (the hook operon, a basal body gene flbN, and the flaO gene), we have measured the level of chimeric gene expression in 13 flagellar mutant backgrounds. Mutants in the hook operon or in basal body genes caused overproduction of both hook operon and basal body gene chimeric mRNAs, suggesting that negative regulation is superimposed on the positive trans-acting control for these early events in the flagellar hierarchy. Mutants in the structural genes and in genes involved in flagellar assembly had no effect on flaO expression, placing the flaO gene near the top of the hierarchy. However, flaO expression appears to be under negative control by two regulatory genes flaS and flaW. Negative control, as a response to the completion of specific steps in the assembly process, may be an important mechanism used by the cell to turn off flagellar gene expression once the gene product is no longer needed.

scholarly journals Multiple Promoters Contribute to Swarming and the Coordination of Transcription with Flagellar Assembly in Salmonella

2010 ◽  
Vol 192 (18) ◽  
pp. 4752-4762 ◽  
Author(s):  
Christopher E. Wozniak ◽  
Fabienne F. V. Chevance ◽  
Kelly T. Hughes
Keyword(s):  
Transcriptional Regulation ◽  
Stationary Phase ◽  
Basal Body ◽  
Environmental Conditions ◽  
Swarming Motility ◽  
Multiple Promoters ◽  
Associated Proteins ◽  
Flagellar Assembly

ABSTRACT In Salmonella, there are three classes of promoters in the flagellar transcriptional hierarchy. This organization allows genes needed earlier in the construction of flagella to be transcribed before genes needed later. Four operons (fliAZY, flgMN, fliDST, and flgKL) are expressed from both class 2 and class 3 promoters. To investigate the purpose for expressing genes from multiple flagellar promoters, mutants were constructed for each operon that were defective in either class 2 transcription or class 3 transcription. The mutants were checked for defects in swimming through liquids, swarming over surfaces, and transcriptional regulation. The expression of the hook-associated proteins (FlgK, FlgL, and FliD) from class 3 promoters was found to be important for swarming motility. Both flgMN promoters were involved in coordinating class 3 transcription with the stage of assembly of the hook-basal body. Finally, the fliAZY class 3 promoter lowered class 3 transcription in stationary phase. These results indicate that the multiple flagellar promoters respond to specific environmental conditions and help coordinate transcription with flagellar assembly.

scholarly journals Bacterial Flagellar Filament: A Supramolecular Multifunctional Nanostructure

2021 ◽  
Vol 22 (14) ◽  
pp. 7521
Author(s):  
Marko Nedeljković ◽  
Diego Emiliano Sastre ◽  
Eric John Sundberg
Keyword(s):  
Immune Responses ◽  
Basal Body ◽  
Vaccine Development ◽  
Bacterial Species ◽  
High Variability ◽  
Bacterial Survival ◽  
Bacterial Flagellum ◽  
Capping Proteins ◽  
Flagellar Filament ◽  
Flagellar Assembly

The bacterial flagellum is a complex and dynamic nanomachine that propels bacteria through liquids. It consists of a basal body, a hook, and a long filament. The flagellar filament is composed of thousands of copies of the protein flagellin (FliC) arranged helically and ending with a filament cap composed of an oligomer of the protein FliD. The overall structure of the filament core is preserved across bacterial species, while the outer domains exhibit high variability, and in some cases are even completely absent. Flagellar assembly is a complex and energetically costly process triggered by environmental stimuli and, accordingly, highly regulated on transcriptional, translational and post-translational levels. Apart from its role in locomotion, the filament is critically important in several other aspects of bacterial survival, reproduction and pathogenicity, such as adhesion to surfaces, secretion of virulence factors and formation of biofilms. Additionally, due to its ability to provoke potent immune responses, flagellins have a role as adjuvants in vaccine development. In this review, we summarize the latest knowledge on the structure of flagellins, capping proteins and filaments, as well as their regulation and role during the colonization and infection of the host.

Pax1 is expressed during development of the thymus epithelium and is required for normal T-cell maturation

Development ◽  
1996 ◽  
Vol 122 (1) ◽  
pp. 23-30 ◽  
Author(s):  
J. Wallin ◽  
H. Eibel ◽  
A. Neubuser ◽  
J. Wilting ◽  
H. Koseki ◽  
...  
Keyword(s):  
T Cell ◽  
Regulatory Protein ◽  
Cell Maturation ◽  
Thymic Epithelial Cells ◽  
Total Size ◽  
Thymic Development ◽  
Transcriptional Regulatory ◽  
Thymus Cells ◽  
T Cell Maturation ◽  
Third Pharyngeal Pouch

Pax1 is a transcriptional regulatory protein expressed during mouse embryogenesis and has been shown to have an important function in vertebral column development. Expression of Pax1 mRNA in the embryonic thymus has been reported previously. Here we show that Pax1 protein expression in thymic epithelial cells can be detected throughout thymic development and in the adult. Expression starts in the early endodermal epithelium lining the foregut region and includes the epithelium of the third pharyngeal pouch, a structure giving rise to part of the thymus epithelium. In early stages of thymus development a large proportion of thymus cells expresses Pax1. With increasing age, the proportion of Pax1-expressing cells is reduced and in the adult mouse only a small fraction of cortical thymic stromal cells retains strong Pax1 expression. Expression of Pax1 in thymus epithelium is necessary for establishing the thymus microenvironment required for normal T cell maturation. Mutations in the Pax-1 gene in undulated mice affect not only the total size of the thymus but also the maturation of thymocytes. The number of thymocytes is reduced about 2- to 5-fold, affecting mainly the CD4+8+ immature and CD4+ mature thymocyte subsets. The expression levels of major thymocyte surface markers remains unchanged with the exception of Thy-1 which was found to be expressed at 3- to 4-fold higher levels.

scholarly journals Helicobacter pylori FlhA Binds the Sensor Kinase and Flagellar Gene Regulatory Protein FlgS with High Affinity

2015 ◽  
Vol 197 (11) ◽  
pp. 1886-1892 ◽  
Author(s):  
Jennifer Tsang ◽  
Takanori Hirano ◽  
Timothy R. Hoover ◽  
Jonathan L. McMurry
Keyword(s):  
Helicobacter Pylori ◽  
Response Regulator ◽  
Regulatory Protein ◽  
Kinase Domain ◽  
Optical Biosensing ◽  
Content Type ◽  
High Affinity ◽  
Flagellar Assembly ◽  
Flagellar Biogenesis

ABSTRACTFlagellar biogenesis is a complex process that involves multiple checkpoints to coordinate transcription of flagellar genes with the assembly of the flagellum. InHelicobacter pylori, transcription of the genes needed in the middle stage of flagellar biogenesis is governed by RpoN and the two-component system consisting of the histidine kinase FlgS and response regulator FlgR. In response to an unknown signal, FlgS autophosphorylates and transfers the phosphate to FlgR, initiating transcription from RpoN-dependent promoters. In the present study, export apparatus protein FlhA was examined as a potential signal protein. Deletion of its N-terminal cytoplasmic sequence dramatically decreased expression of two RpoN-dependent genes,flaBandflgE. Optical biosensing demonstrated a high-affinity interaction between FlgS and a peptide consisting of residues 1 to 25 of FlhA (FlhANT). TheKD(equilibrium dissociation constant) was 21 nM and was characterized by fast-on (kon= 2.9 × 104M−1s−1) and slow-off (koff= 6.2 × 10−4s−1) kinetics. FlgS did not bind peptides consisting of smaller fragments of the FlhANTsequence. Analysis of binding to purified fragments of FlgS demonstrated that the C-terminal portion of the protein containing the kinase domain binds FlhANT. FlhANTbinding did not stimulate FlgS autophosphorylationin vitro, suggesting that FlhA facilitates interactions between FlgS and other structures required to stimulate autophosphorylation.IMPORTANCEThe high-affinity binding of FlgS to FlhA characterized in this study points to an additional role for FlhA in flagellar assembly. Beyond its necessity for type III secretion, the N-terminal cytoplasmic sequence of FlhA is required for RpoN-dependent gene expression via interaction with the C-terminal kinase domain of FlgS.

Control of beta-interferon expression in murine embryonal carcinoma F9 cells

1989 ◽  
Vol 9 (8) ◽  
pp. 3553-3556
Author(s):  
M K Francis ◽  
J M Lehman
Keyword(s):  
Retinoic Acid ◽  
Embryonal Carcinoma ◽  
Biologically Active ◽  
Rnase Protection ◽  
F9 Cells ◽  
Differentiated Cells ◽  
Transcriptional Regulatory Mechanism ◽  
Transcriptional Regulatory ◽  
Tissue Culture Model

Murine embryonal carcinoma F9 cells, a tissue culture model for early embryonic development, do not produce interferon (IFN) in response to poly(I-C), as determined by an antiviral assay. RNase protection analyses were used to examine total RNA extracted from the cells for the presence of beta-IFN RNA. Whereas F9 cells differentiated in vitro with retinoic acid produced a biologically active protein as well as beta-IFN RNA in response to poly(I-C), undifferentiated F9 cells produced no detectable beta-IFN RNA even in the presence of cycloheximide, an IFN-superinducing agent. These results show that undifferentiated embryonal carcinoma cells do not accumulate beta-IFN RNA in response to an IFN-inducing agent, suggesting a transcriptional regulatory mechanism. However, this control mechanism is altered upon differentiation, since the gene can be transcriptionally activated in retinoic acid-differentiated cells.

scholarly journals In SituStructures of Polar and Lateral Flagella Revealed by Cryo-Electron Tomography

2019 ◽  
Vol 201 (13) ◽  
Author(s):  
Shiwei Zhu ◽  
Maren Schniederberend ◽  
Daniel Zhitnitsky ◽  
Ruchi Jain ◽  
Jorge E. Galán ◽  
...  
Keyword(s):  
Salmonella Enterica ◽  
Electron Tomography ◽  
Additional Insight ◽  
Content Type ◽  
Bacterial Flagellum ◽  
Cryo Electron Tomography ◽  
Serovar Typhimurium ◽  
Flagellar Assembly ◽  
Insight Into

ABSTRACTThe bacterial flagellum is a sophisticated self-assembling nanomachine responsible for motility in many bacterial pathogens, includingPseudomonas aeruginosa,Vibriospp., andSalmonella enterica. The bacterial flagellum has been studied extensively in the model systemsEscherichia coliandSalmonella entericaserovar Typhimurium, yet the range of variation in flagellar structure and assembly remains incompletely understood. Here, we used cryo-electron tomography and subtomogram averaging to determinein situstructures of polar flagella inP. aeruginosaand peritrichous flagella inS. Typhimurium, revealing notable differences between these two flagellar systems. Furthermore, we observed flagellar outer membrane complexes as well as many incomplete flagellar subassemblies, which provide additional insight into mechanisms underlying flagellar assembly and loss in bothP. aeruginosaandS. Typhimurium.IMPORTANCEThe bacterial flagellum has evolved as one of the most sophisticated self-assembled molecular machines, which confers locomotion and is often associated with virulence of bacterial pathogens. Variation in species-specific features of the flagellum, as well as in flagellar number and placement, results in structurally distinct flagella that appear to be adapted to the specific environments that bacteria encounter. Here, we used cutting-edge imaging techniques to determine high-resolutionin situstructures of polar flagella inPseudomonas aeruginosaand peritrichous flagella inSalmonella entericaserovar Typhimurium, demonstrating substantial variation between flagella in these organisms. Importantly, we observed novel flagellar subassemblies and provided additional insight into the structural basis of flagellar assembly and loss in bothP. aeruginosaandS. Typhimurium.

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