scholarly journals Diversification of Campylobacter jejuni Flagellar C-Ring Composition Impacts Its Structure and Function in Motility, Flagellar Assembly, and Cellular Processes

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Louie D. Henderson ◽  
Teige R. S. Matthews-Palmer ◽  
Connor J. Gulbronson ◽  
Deborah A. Ribardo ◽  
Morgan Beeby ◽  
...  
Keyword(s):  
Campylobacter Jejuni ◽  
Content Type ◽  
Cellular Processes ◽  
Spatial Regulation ◽  
Flagellar Motors ◽  
Core Components ◽  
Flagellar Assembly ◽  
And Function ◽  
Conserved Core ◽  
Flagellar Biogenesis

ABSTRACT Bacterial flagella are reversible rotary motors that rotate external filaments for bacterial propulsion. Some flagellar motors have diversified by recruiting additional components that influence torque and rotation, but little is known about the possible diversification and evolution of core motor components. The mechanistic core of flagella is the cytoplasmic C ring, which functions as a rotor, directional switch, and assembly platform for the flagellar type III secretion system (fT3SS) ATPase. The C ring is composed of a ring of FliG proteins and a helical ring of surface presentation of antigen (SPOA) domains from the switch proteins FliM and one of two usually mutually exclusive paralogs, FliN or FliY. We investigated the composition, architecture, and function of the C ring of Campylobacter jejuni, which encodes FliG, FliM, and both FliY and FliN by a variety of interrogative approaches. We discovered a diversified C. jejuni C ring containing FliG, FliM, and both FliY, which functions as a classical FliN-like protein for flagellar assembly, and FliN, which has neofunctionalized into a structural role. Specific protein interactions drive the formation of a more complex heterooligomeric C. jejuni C-ring structure. We discovered that this complex C ring has additional cellular functions in polarly localizing FlhG for numerical regulation of flagellar biogenesis and spatial regulation of division. Furthermore, mutation of the C. jejuni C ring revealed a T3SS that was less dependent on its ATPase complex for assembly than were other systems. Our results highlight considerable evolved flagellar diversity that impacts motor output, biogenesis, and cellular processes in different species. IMPORTANCE The conserved core of bacterial flagellar motors reflects a shared evolutionary history that preserves the mechanisms essential for flagellar assembly, rotation, and directional switching. In this work, we describe an expanded and diversified set of core components in the Campylobacter jejuni flagellar C ring, the mechanistic core of the motor. Our work provides insight into how usually conserved core components may have diversified by gene duplication, enabling a division of labor of the ancestral protein between the two new proteins, acquisition of new roles in flagellar assembly and motility, and expansion of the function of the flagellum beyond motility, including spatial regulation of cell division and numerical control of flagellar biogenesis in C. jejuni. Our results highlight that relatively small changes, such as gene duplications, can have substantial ramifications on the cellular roles of a molecular machine.

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.

scholarly journals Campylobacter jejuni Triggers Signaling through Host Cell Focal Adhesions To Inhibit Cell Motility

mBio ◽  
2021 ◽  
Author(s):  
Courtney M. Klappenbach ◽  
Nicholas M. Negretti ◽  
Jesse Aaron ◽  
Teng-Leong Chew ◽  
Michael E. Konkel
Keyword(s):  
Epithelial Cells ◽  
Cell Motility ◽  
Host Cell ◽  
Campylobacter Jejuni ◽  
Focal Adhesion ◽  
Foodborne Pathogen ◽  
Focal Adhesions ◽  
Structure And Function ◽  
Content Type ◽  
And Function

Campylobacter jejuni is a major foodborne pathogen that causes severe gastritis. We investigated the dynamics of focal adhesion structure and function in C. jejuni -infected epithelial cells.

scholarly journals Evidence of Intraflagellar Transport and Apical Complex Formation in a Free-Living Relative of the Apicomplexa

2013 ◽  
Vol 13 (1) ◽  
pp. 10-20 ◽  
Author(s):  
Neil Portman ◽  
Christie Foster ◽  
Giselle Walker ◽  
Jan Šlapeta
Keyword(s):  
Close Association ◽  
Flagellar Apparatus ◽  
Intraflagellar Transport ◽  
Endomembrane System ◽  
Free Living ◽  
Content Type ◽  
Daughter Cells ◽  
Flagellar Assembly ◽  
Apical Complex ◽  
Flagellar Biogenesis

ABSTRACT Since its first description, Chromera velia has attracted keen interest as the closest free-living relative of parasitic Apicomplexa. The life cycle of this unicellular alga is complex and involves a motile biflagellate form. Flagella are thought to be formed in the cytoplasm, a rare phenomenon shared with Plasmodium in which the canonical mode of flagellar assembly, intraflagellar transport, is dispensed with. Here we demonstrate the expression of intraflagellar transport components in C. velia , answering the question of whether this organism has the potential to assemble flagella via the canonical route. We have developed and characterized a culturing protocol that favors the generation of flagellate forms. From this, we have determined a marked shift in the mode of daughter cell production from two to four daughter cells per division as a function of time after passage. We conduct an ultrastructural examination of the C. velia flagellate form by using serial TEM and show that flagellar biogenesis in C. velia occurs prior to cytokinesis. We demonstrate a close association of the flagellar apparatus with a complex system of apical structures, including a micropore, a conoid, and a complex endomembrane system reminiscent of the apical complex of parasitic apicomplexans. Recent work has begun to elucidate the possible flagellar origins of the apical complex, and we show that in C. velia these structures are contemporaneous within a single cell and share multiple connections. We propose that C. velia therefore represents a vital piece in the puzzle of the origins of the apical complex.

scholarly journals Imaging the Motility and Chemotaxis Machineries in Helicobacter pylori by Cryo-Electron Tomography

2016 ◽  
Vol 199 (3) ◽  
Author(s):  
Zhuan Qin ◽  
Wei-ting Lin ◽  
Shiwei Zhu ◽  
Aime T. Franco ◽  
Jun Liu
Keyword(s):  
Helicobacter Pylori ◽  
Molecular Mechanisms ◽  
Electron Tomography ◽  
Bacterial Pathogen ◽  
Human Stomach ◽  
Content Type ◽  
Flagellar Motors ◽  
Cryo Electron Tomography ◽  
Flagellar Assembly ◽  
H Pylori

ABSTRACT Helicobacter pylori is a bacterial pathogen that can cause many gastrointestinal diseases, including ulcers and gastric cancer. A unique chemotaxis-mediated motility is critical for H. pylori to colonize in the human stomach and to establish chronic infection, but the underlying molecular mechanisms are not well understood. Here, we employ cryo-electron tomography (cryo-ET) to reveal detailed structures of the H. pylori cell envelope, including the sheathed flagella and chemotaxis arrays. Notably, H. pylori possesses a distinctive periplasmic cage-like structure with 18-fold symmetry. We propose that this structure forms a robust platform for recruiting 18 torque generators, which likely provide the higher torque needed for swimming in high-viscosity environments. We also reveal a series of key flagellar assembly intermediates, providing structural evidence that flagellar assembly is tightly coupled with the biogenesis of the membrane sheath. Finally, we determine the structure of putative chemotaxis arrays at the flagellar pole, which have implications for how the direction of flagellar rotation is regulated. Together, our pilot cryo-ET studies provide novel structural insights into the unipolar flagella of H. pylori and lay a foundation for a better understanding of the unique motility of this organism. IMPORTANCE Helicobacter pylori is a highly motile bacterial pathogen that colonizes approximately 50% of the world's population. H. pylori can move readily within the viscous mucosal layer of the stomach. It has become increasingly clear that its unique flagella-driven motility is essential for successful gastric colonization and pathogenesis. Here, we use advanced imaging techniques to visualize novel in situ structures with unprecedented detail in intact H. pylori cells. Remarkably, H. pylori possesses multiple unipolar flagella, which are driven by one of the largest flagellar motors found in bacteria. These large motors presumably provide the higher torque needed by the bacterial pathogens to navigate in the viscous environment of the human stomach.

scholarly journals A Regulatory Checkpoint during Flagellar Biogenesis in Campylobacter jejuni Initiates Signal Transduction To Activate Transcription of Flagellar Genes

mBio ◽  
2013 ◽  
Vol 4 (5) ◽  
Author(s):  
Joseph M. Boll ◽  
David R. Hendrixson
Keyword(s):  
Signal Transduction ◽  
Campylobacter Jejuni ◽  
Gene Transcription ◽  
Type Iii Secretion ◽  
Secretion System ◽  
Flagellar Gene ◽  
Content Type ◽  
Regulatory Systems ◽  
Two Component ◽  
Flagellar Biogenesis

ABSTRACTMany polarly flagellated bacteria require similar two-component regulatory systems (TCSs) and σ54to activate transcription of genes essential for flagellar motility. Herein, we discovered that in addition to the flagellar type III secretion system (T3SS), theCampylobacter jejuniflagellar MS ring and rotor are required to activate the FlgSR TCS. Mutants lacking the FliF MS ring and FliG C ring rotor proteins were as defective as T3SS mutants in FlgSR- and σ54-dependent flagellar gene expression. Also, FliF and FliG required each other for stability, which is mediated by atypical extensions to the proteins. A FliF mutant that presumably does not interact with the T3SS protein FlhA did not support flagellar gene transcription, suggesting that FliF-T3SS interactions are essential to generate a signal sensed by the cytoplasmic FlgS histidine kinase. Furthermore, the flagellar T3SS was required for FlgS to immunoprecipitate with FliF and FliG. We propose a model whereby the flagellar T3SS facilitates FliF and FliG multimerization into the MS ring and rotor. As a result, these flagellar structures form a cytoplasmic complex that interacts with and is sensed by FlgS. The synthesis of these structures appears to be a regulatory checkpoint in flagellar biogenesis that the FlgS kinase monitors to initiate signal transduction that activates σ54and expression of genes required for the next stage of flagellation. Given that other polar flagellates have flagellar transcriptional hierarchies that are organized similarly as inC. jejuni, this regulatory checkpoint may exist in a broad range of bacteria to influence similar TCSs and flagellar gene transcription.IMPORTANCEDespite the presence of numerous two-component regulatory systems (TCSs) in bacteria, direct signals sensed by TCSs to activate signal transduction are known for only a minority. Polar flagellates, includingPseudomonas,Vibrio,Helicobacter, andCampylobacterspecies, require a similar TCS and σ54for flagellar gene transcription, but the activating signals for these TCSs are unknown. We explored signals that activate theCampylobacter jejuniFlgSR TCS to initiate σ54-dependent flagellar gene transcription. Our discoveries suggest that the FlgS histidine kinase monitors the formation of the flagellar type III secretion system and the surrounding MS and C rings. The synthesis of these structures creates a regulatory checkpoint in flagellar biogenesis that is sensed by FlgS to ensure proper transcription of the next set of genes for subsequent steps in flagellation. Given the conservation of flagellar-associated TCSs and transcriptional cascades in polar flagellates, this regulatory checkpoint in flagellar biogenesis likely impacts flagellation in a broad range of bacteria.

scholarly journals RNA Interference in Fungi: Pathways, Functions, and Applications

2011 ◽  
Vol 10 (9) ◽  
pp. 1148-1155 ◽  
Author(s):  
Yunkun Dang ◽  
Qiuying Yang ◽  
Zhihong Xue ◽  
Yi Liu
Keyword(s):  
Gene Regulation ◽  
Rna Interference ◽  
Small Rna ◽  
Rnai Machinery ◽  
Content Type ◽  
Heterochromatin Formation ◽  
Rna Molecules ◽  
Cellular Processes ◽  
Rna Biogenesis ◽  
Core Components

ABSTRACT Small RNA molecules of about 20 to 30 nucleotides function in gene regulation and genomic defense via conserved eukaryotic RNA interference (RNAi)-related pathways. The RNAi machinery consists of three core components: Dicer, Argonaute, and RNA-dependent RNA polymerase. In fungi, the RNAi-related pathways have three major functions: genomic defense, heterochromatin formation, and gene regulation. Studies of Schizosaccharomyces pombe and Neurospora , and other fungi have uncovered surprisingly diverse small RNA biogenesis pathways, suggesting that fungi utilize RNAi-related pathways in various cellular processes to adapt to different environmental conditions. These studies also provided important insights into how RNAi functions in eukaryotic systems in general. In this review, we will discuss our current understanding of the fungal RNAi-related pathways and their functions, with a focus on filamentous fungi. We will also discuss how RNAi can be used as a tool in fungal research.

scholarly journals Central Carbon Metabolism and Electron Transport in Chlamydomonas reinhardtii: Metabolic Constraints for Carbon Partitioning between Oil and Starch

2013 ◽  
Vol 12 (6) ◽  
pp. 776-793 ◽  
Author(s):  
Xenie Johnson ◽  
Jean Alric
Keyword(s):  
Chlamydomonas Reinhardtii ◽  
Carbon Metabolism ◽  
Lipid Production ◽  
Central Carbon Metabolism ◽  
Content Type ◽  
Cellular Processes ◽  
Renewable Feedstock ◽  
Central Carbon ◽  
Redox Poise ◽  
And Function

ABSTRACT The metabolism of microalgae is so flexible that it is not an easy task to give a comprehensive description of the interplay between the various metabolic pathways. There are, however, constraints that govern central carbon metabolism in Chlamydomonas reinhardtii that are revealed by the compartmentalization and regulation of the pathways and their relation to key cellular processes such as cell motility, division, carbon uptake and partitioning, external and internal rhythms, and nutrient stress. Both photosynthetic and mitochondrial electron transfer provide energy for metabolic processes and how energy transfer impacts metabolism and vice versa is a means of exploring the regulation and function of these pathways. A key example is the specific chloroplast localization of glycolysis/gluconeogenesis and how it impacts the redox poise and ATP budget of the plastid in the dark. To compare starch and lipids as carbon reserves, their value can be calculated in terms of NAD(P)H and ATP. As microalgae are now considered a potential renewable feedstock, we examine current work on the subject and also explore the possibility of rerouting metabolism toward lipid production.

scholarly journals A Chaperone for the Stator Units of a Bacterial Flagellum

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Deborah A. Ribardo ◽  
Brittni R. Kelley ◽  
Jeremiah G. Johnson ◽  
David R. Hendrixson
Keyword(s):  
Campylobacter Jejuni ◽  
Bacterial Species ◽  
Evolutionary Adaptation ◽  
Flagellar Motor ◽  
Content Type ◽  
Bacterial Flagellum ◽  
Flagellar Motors ◽  
Sufficient Power ◽  
Ftsh Protease ◽  
Flagellar Rotation

ABSTRACT The stator units of the flagellum supply power to the flagellar motor via ion transport across the cytoplasmic membrane and generate torque on the rotor for rotation. Flagellar motors across bacterial species have evolved adaptations that impact and enhance stator function to meet the demands of each species, including producing stator units using different fuel types or various stator units for different motility modalities. Campylobacter jejuni produces one of the most complex and powerful flagellar motors by positioning 17 stator units at a greater radial distance than in most other bacteria to increase power and torque for high velocity of motility. We report another evolutionary adaptation impacting flagellar stators by identifying FlgX as a chaperone for C. jejuni stator units to ensure sufficient power and torque for flagellar rotation and motility. We discovered that FlgX maintains MotA and MotB stator protein integrity likely through a direct interaction with MotA that prevents their degradation. Suppressor analysis suggested that the physiology of C. jejuni drives the requirement for FlgX to protect stator units from proteolysis by the FtsH protease complex. C. jejuni ΔflgX was strongly attenuated for colonization of the natural avian host, but colonization capacity was greatly restored by a single mutation in MotA. These findings suggest that the likely sole function of FlgX is to preserve stator unit integrity for the motility required for host interactions. Our findings demonstrate another evolved adaptation in motile bacteria to ensure the equipment of the flagellar motor with sufficient power to generate torque for motility. IMPORTANCE The bacterial flagellum is a reversible rotating motor powered by ion transport through stator units, which also exert torque on the rotor component to turn the flagellum for motility. Species-specific adaptations to flagellar motors impact stator function to meet the demands of each species to sufficiently power flagellar rotation. We identified another evolutionary adaptation by discovering that FlgX of Campylobacter jejuni preserves the integrity of stator units by functioning as a chaperone to protect stator proteins from degradation by the FtsH protease complex due to the physiology of the bacterium. FlgX is required to maintain a level of stator units sufficient to power the naturally high-torque flagellar motor of C. jejuni for motility in intestinal mucosal layers to colonize hosts. Our work continues to identify an increasing number of adaptations to flagellar motors across bacterial species that provide the mechanics necessary for producing an effective rotating nanomachine for motility.

scholarly journals Under Elevated c-di-GMP in Escherichia coli, YcgR Alters Flagellar Motor Bias and Speed Sequentially, with Additional Negative Control of the Flagellar Regulon via the Adaptor Protein RssB

2019 ◽  
Vol 202 (1) ◽  
Author(s):  
Vincent Nieto ◽  
Jonathan D. Partridge ◽  
Geoffrey B. Severin ◽  
Run-Zhi Lai ◽  
Christopher M. Waters ◽  
...  
Keyword(s):  
Adaptor Protein ◽  
Negative Control ◽  
Flagellar Motor ◽  
Content Type ◽  
Environmental Signals ◽  
E Coli ◽  
Regulator Protein ◽  
Flagellar Regulon ◽  
Flagellar Assembly ◽  
And Function

Flagellum-driven motility has been studied in E. coli and Salmonella for nearly half a century. Over 60 genes control flagellar assembly and function. The expression of these genes is regulated at multiple levels in response to a variety of environmental signals. Cues that elevate c-di-GMP levels, however, inhibit motility by direct binding of the effector YcgR to the flagellar motor. In this study conducted mainly in E. coli, we show that YcgR is the only effector of motor control and tease out the order of YcgR-mediated events. In addition, we find that the σS regulator protein RssB contributes to negative regulation of flagellar gene expression when c-di-GMP levels are elevated.

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