Dynamics of the two stator systems in the flagellar motor of Pseudomonas aeruginosa studied by a bead assay

2021 ◽  
Author(s):  
Zhengyu Wu ◽  
Maojin Tian ◽  
Rongjing Zhang ◽  
Junhua Yuan
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Single Cell ◽  
Motor Behavior ◽  
Optimal Performance ◽  
Polar Flagellum ◽  
Flagellar Motor ◽  
Motor Speed ◽  
Wild Type ◽  
Flagellar Filament ◽  
Environmental Exploration

We developed a robust bead assay for studying flagellar motor behavior of Pseudomonas aeruginosa . Using this assay, we studied the dynamics of the two stator systems in the flagellar motor. We found that the two sets of stators function differently, with MotAB stators providing higher total torque, and MotCD stators ensuring more stable motor speed. The motors in wild-type cells adjust the stator compositions according to the environment, resulting in an optimal performance in environmental exploration compared to mutants with one set of stators. The bead assay we developed here can be further used to study P. aeruginosa chemotaxis at the level of single cell using the motor behavior as the chemotaxis output. Importance Cells of Pseudomonas aeruginosa possess a single polar flagellum, driven by a rotatory motor powered by two sets of torque-generating units (stators). We developed a robust bead assay for studying the behavior of the flagellar motor in P. aeruginosa , by attaching a microsphere to shortened flagellar filament and using it as an indicator of motor rotation. Using this assay, we revealed the dynamics of the two stator systems in the flagellar motor, and found that the motors in wild-type cells adjust the stator compositions according to the environment, resulting in an optimal performance in environmental exploration compared to mutants with one set of stators.

scholarly journals Characterization of chemotaxis and motility response towards fructose in Escherichia coli

2020 ◽  
Author(s):  
Fanghai Liu ◽  
Fangbin Wang ◽  
Jian Liu
Keyword(s):  
Escherichia Coli ◽  
Motor Behavior ◽  
Stimulus Level ◽  
Flagellar Motor ◽  
Motor Speed ◽  
Wild Type ◽  
Biased Random Walk ◽  
Bacterial Aggregation ◽  
Chemotaxis System

Abstract BackgroundPeritrichously flagellated bacteria such as Escherichia coli perform chemotaxis by a biased random walk toward various chemicals, which was driven by the bacterial flagellar motor. The chemotaxis system is a perfect robust system which can adapt to its pre-stimulus level after the stimulus. Fructose, a typical monosaccharide that can attract Escherichia coli. Previous studies have shown Escherichia coli to be chemotactic to both amino acids and simple sugars. However, little is known about the chemotaxis and motility response of Escherichia coli towards fructose.ResultsIn this study, we characterized the chemotaxis behavior of Escherichia coli to different concentrations of fructose from 0mM to 50mM by using microfluidics and bead assay. We observed the wild-type cells responded to the stimulus of fructose, which suggested fructose is an attractant to Escherichia coli, while the cells defective in chemotaxis could not sense the stimulus of fructose. Fructose affects the motor behavior of wild-type cells and reduces the CCW motor speeds significantly compared to CW motor speeds, contrast to that fructose cannot affect the motor speeds of cells defective in chemotaxis. The CW biases of wild-type cells increased with the addition of fructose. And the fructose promotes bacterial aggregation on surface by lowering the bacteria motor speed.ConclusionsWe found that fructose is an attractant to Escherichia coli, with the wild-type cells showed stronger response to higher concentration of fructose, while cells lacking chemoreceptors and CheY cannot sense or respond to fructose. The motility of wild-type cells was reduced in various concentrations of fructose, which helped the aggregation of cells near the surface, in contrast with the result that the fructose showed no effect on the motility of the cells defective in chemotaxis. Similar phenomena are expected to be found in the effect of other monosaccharides to Escherichia coli.

scholarly journals Structural insights into flagellar stator–rotor interactions

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Yunjie Chang ◽  
Ki Hwan Moon ◽  
Xiaowei Zhao ◽  
Steven J Norris ◽  
MD A Motaleb ◽  
...  
Keyword(s):  
Conformational Change ◽  
High Speed ◽  
Electron Tomography ◽  
Deletion Mutants ◽  
Flagellar Motor ◽  
Wild Type ◽  
Cryo Electron Tomography ◽  
Flagellar Filament ◽  
Structural Insights

The bacterial flagellar motor is a molecular machine that can rotate the flagellar filament at high speed. The rotation is generated by the stator–rotor interaction, coupled with an ion flux through the torque-generating stator. Here we employed cryo-electron tomography to visualize the intact flagellar motor in the Lyme disease spirochete, Borrelia burgdorferi. By analyzing the motor structures of wild-type and stator-deletion mutants, we not only localized the stator complex in situ, but also revealed the stator–rotor interaction at an unprecedented detail. Importantly, the stator–rotor interaction induces a conformational change in the flagella C-ring. Given our observation that a non-motile mutant, in which proton flux is blocked, cannot generate the similar conformational change, we propose that the proton-driven torque is responsible for the conformational change required for flagellar rotation.

scholarly journals Single Cells Exhibit Differing Behavioral Phases during Early Stages of Pseudomonas aeruginosa Swarming

2019 ◽  
Vol 201 (19) ◽  
Author(s):  
Chinedu S. Madukoma ◽  
Peixian Liang ◽  
Aleksandar Dimkovikj ◽  
Jianxu Chen ◽  
Shaun W. Lee ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Cell Motility ◽  
Single Cell ◽  
High Cell Density ◽  
Single Cells ◽  
High Cell ◽  
Wild Type ◽  
Content Type ◽  
Type Iv ◽  
The Many

ABSTRACT Pseudomonas aeruginosa is among the many bacteria that swarm, where groups of cells coordinate to move over surfaces. It has been challenging to determine the behavior of single cells within these high-cell-density swarms. To track individual cells within P. aeruginosa swarms, we imaged a fluorescently labeled subset of the larger population. Single cells at the advancing swarm edge varied in their motility dynamics as a function of time. From these data, we delineated four phases of early swarming prior to the formation of the tendril fractals characteristic of P. aeruginosa swarming by collectively considering both micro- and macroscale data. We determined that the period of greatest single-cell motility does not coincide with the period of greatest collective swarm expansion. We also noted that flagellar, rhamnolipid, and type IV pilus motility mutants exhibit substantially less single-cell motility than the wild type. IMPORTANCE Numerous bacteria exhibit coordinated swarming motion over surfaces. It is often challenging to assess the behavior of single cells within swarming communities due to the limitations of identifying, tracking, and analyzing the traits of swarming cells over time. Here, we show that the behavior of Pseudomonas aeruginosa swarming cells can vary substantially in the earliest phases of swarming. This is important to establish that dynamic behaviors should not be assumed to be constant over long periods when predicting and simulating the actions of swarming bacteria.

scholarly journals The Complex Flagellar Torque Generator of Pseudomonas aeruginosa

2004 ◽  
Vol 186 (19) ◽  
pp. 6341-6350 ◽  
Author(s):  
Timothy B. Doyle ◽  
Andrew C. Hawkins ◽  
Linda L. McCarter
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Membrane Proteins ◽  
Motor Function ◽  
Swimming Speed ◽  
Flagellar Motor ◽  
Loss Of Function ◽  
Wild Type ◽  
Aqueous Environments ◽  
Genes Encoding ◽  
Rotary Motors

ABSTRACT Flagella act as semirigid helical propellers that are powered by reversible rotary motors. Two membrane proteins, MotA and MotB, function as a complex that acts as the stator and generates the torque that drives rotation. The genome sequence of Pseudomonas aeruginosa PAO1 contains dual sets of motA and motB genes, PA1460-PA1461 (motAB) and PA4954-PA4953 (motCD), as well as another gene, motY (PA3526), which is known to be required for motor function in some bacteria. Here, we show that these five genes contribute to motility. Loss of function of either motAB-like locus was dispensable for translocation in aqueous environments. However, swimming could be entirely eliminated by introduction of combinations of mutations in the two motAB-encoding regions. Mutation of both genes encoding the MotA homologs or MotB homologs was sufficient to abolish motility. Mutants carrying double mutations in nonequivalent genes (i.e., motA motD or motB motC) retained motility, indicating that noncognate components can function together. motY appears to be required for motAB function. The combination of motY and motCD mutations rendered the cells nonmotile. Loss of function of motAB, motY, or motAB motY produced similar phenotypes; although the swimming speed was only reduced to ∼85% of the wild-type speed, translocation in semisolid motility agar and swarming on the surface of solidified agar were severely impeded. Thus, the flagellar motor of P. aeruginosa represents a more complex configuration than the configuration that has been studied in other bacteria, and it enables efficient movement under different circumstances.

scholarly journals Aeromonas hydrophila motY is essential for polar flagellum function, and requires coordinate expression of motX and Pom proteins

Microbiology ◽  
2011 ◽  
Vol 157 (10) ◽  
pp. 2772-2784 ◽  
Author(s):  
Raquel Molero ◽  
Markus Wilhelms ◽  
Belén Infanzón ◽  
Juan M. Tomás ◽  
Susana Merino
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Aeromonas Hydrophila ◽  
Vibrio Parahaemolyticus ◽  
Proton Motive Force ◽  
Electrochemical Potential ◽  
Polar Flagellum ◽  
Wild Type ◽  
Swimming Motility ◽  
Essential Proteins ◽  
Coordinate Expression

By the analysis of the Aeromonas hydrophila ATCC7966T genome we identified A. hydrophila AH-3 MotY. A. hydrophila MotY, like MotX, is essential for the polar flagellum function energized by an electrochemical potential of Na+ as coupling ion, but is not involved in lateral flagella function energized by the proton motive force. Thus, the A. hydrophila polar flagellum stator is a complex integrated by two essential proteins, MotX and MotY, which interact with one of two redundant pairs of proteins, PomAB and PomA2B2. In an A. hydrophila motX mutant, polar flagellum motility is restored by motX complementation, but the ability of the A. hydrophila motY mutant to swim is not restored by introduction of the wild-type motY alone. However, its polar flagellum motility is restored when motX and -Y are expressed together from the same plasmid promoter. Finally, even though both the redundant A. hydrophila polar flagellum stators, PomAB and PomA2B2, are energized by the Na+ ion, they cannot be exchanged. Furthermore, Vibrio parahaemolyticus PomAB and Pseudomonas aeruginosa MotAB or MotCD are unable to restore swimming motility in A. hydrophila polar flagellum stator mutants.

scholarly journals Biofilm formation by Pseudomonas aeruginosa wild type, flagella and type IV pili mutants

2003 ◽  
Vol 48 (6) ◽  
pp. 1511-1524 ◽  
Author(s):  
Mikkel Klausen ◽  
Arne Heydorn ◽  
Paula Ragas ◽  
Lotte Lambertsen ◽  
Anders Aaes-Jørgensen ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Biofilm Formation ◽  
Type Iv Pili ◽  
Wild Type ◽  
Type Iv

scholarly journals ParB deficiency in Pseudomonas aeruginosa destabilizes the partner protein ParA and affects a variety of physiological parameters

Microbiology ◽  
2009 ◽  
Vol 155 (4) ◽  
pp. 1080-1092 ◽  
Author(s):  
A. A. Bartosik ◽  
J. Mierzejewska ◽  
C. M. Thomas ◽  
G. Jagura-Burdzy
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Fluorescence Microscopy ◽  
Bacterial Growth ◽  
Complete Loss ◽  
Physiological Parameters ◽  
Wild Type ◽  
Partial Removal ◽  
Partner Protein ◽  
The One ◽  
Mutant Phenotypes

Deletions leading to complete or partial removal of ParB were introduced into the Pseudomonas aeruginosa chromosome. Fluorescence microscopy of fixed cells showed that ParB mutants lacking the C-terminal domain or HTH motif formed multiple, less intense foci scattered irregularly, in contrast to the one to four ParB foci per cell symmetrically distributed in wild-type P. aeruginosa. All parB mutations affected both bacterial growth and swarming and swimming motilities, and increased the production of anucleate cells. Similar effects were observed after inactivation of parA of P. aeruginosa. As complete loss of ParA destabilized its partner ParB it was unclear deficiency of which protein is responsible for the mutant phenotypes. Analysis of four parB mutants showed that complete loss of ParB destabilized ParA whereas three mutants that retained the N-terminal 90 aa of ParB did not. As all four parB mutants demonstrate the same defects it can be concluded that either ParB, or ParA and ParB in combination, plays an important role in nucleoid distribution, growth and motility in P. aeruginosa.

Antibacterial effect of hyaluronan/chitosan nanofilm in the initial adhesion of Pseudomonas aeruginosa wild type, and IV pili and LPS mutant strains

2021 ◽  
pp. 101415
Author(s):  
Jacobo Hernandez-Montelongo ◽  
Gianlucca G. Nicastro ◽  
Thays de O. Pereira ◽  
Mariana Zavarize ◽  
Marisa M. Beppu ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Antibacterial Effect ◽  
Wild Type ◽  
Initial Adhesion ◽  
Mutant Strains

scholarly journals IMMU-35. TRANSCRIPTIONALLY DEFINED IMMUNE CONTEXTURE IN HUMAN GLIOMAS AT SINGLE-CELL RESOLUTION

2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii112-ii112
Author(s):  
Pravesh Gupta ◽  
Minghao Dang ◽  
Krishna Bojja ◽  
Tuan Tran M ◽  
Huma Shehwana ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Single Cell ◽  
Rna Sequencing ◽  
Immune Cell ◽  
Wild Type ◽  
Sequencing Data ◽  
Human Gliomas ◽  
Cell Clusters ◽  
Recurrent Gliomas ◽  
Antigen Presenting

Abstract The brain tumor immune microenvironment (TIME) continuously evolves during glioma progression and a comprehensive understanding of the glioma-centric immune cell repertoire beyond a priori cell types and/or states is uncharted. Consequently, we performed single-cell RNA-sequencing on ~123,000 tumor-derived immune cells from 17-pathologically stratified, IDH (isocitrate dehydrogenase)-differential primary, recurrent human gliomas, and non-glioma brains. Our analysis delineated predominant 34-myeloid cell clusters (~75%) over 28-lymphoid cell clusters (~25%) reflecting enormous heterogeneity within and across gliomas. The glioma immune diversity spanned functionally imprinted phagocytic, antigen-presenting, hypoxia, angiogenesis and, tumoricidal myeloid to classical cytotoxic lymphoid subpopulations. Specifically, IDH-mutant gliomas were enriched for brain-resident microglial subpopulations in contrast to enhanced bone barrow-derived infiltrates in IDH-wild type, especially in a recurrent setting. Microglia attrition in IDH-wild type -primary and -recurrent gliomas were concomitant with invading monocyte-derived cells with semblance to dendritic cell and macrophage/microglia like transcriptomic features. Additionally, microglial functional diversification was noted with disease severity and mostly converged to inflammatory states in IDH-wild type recurrent gliomas. Beyond dendritic cells, multiple antigen-presenting cellular states expanded with glioma severity especially in IDH-wild type primary and recurrent- gliomas. Furthermore, we noted differential microglia and dendritic cell inherent antigen presentation axis viz, osteopontin, and classical HLAs in IDH subtypes and, glioma-wide non-PD1 checkpoints associations in T cells like Galectin9 and Tim-3. As a general utility, our immune cell deconvolution approach with single-cell-matched bulk RNA sequencing data faithfully resolved 58-cell states which provides glioma specific immune reference for digital cytometry application to genomics datasets. Resultantly, we identified prognosticator immune cell-signatures from TCGA cohorts as one of many potential immune responsiveness applications of the curated signatures for basic and translational immune-genomics efforts. Thus, we not only provide an unprecedented insight of glioma TIME but also present an immune data resource that can be exploited to guide pragmatic glioma immunotherapy designs.

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