scholarly journals Competitive binding of independent extension and retraction motors explains the quantitative dynamics of type IV pili

2020 ◽  
Author(s):  
Matthias D. Koch ◽  
Chenyi Fei ◽  
Ned S. Wingreen ◽  
Joshua W. Shaevitz ◽  
Zemer Gitai
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Production Rate ◽  
Opportunistic Pathogen ◽  
Competitive Binding ◽  
Type Iv Pili ◽  
Biophysical Modeling ◽  
Full Extension ◽  
Type Iv ◽  
Surface Contact

AbstractThe functions of type IV pili (TFP) are mediated by cycles of extension and retraction. The coordination of these cycles remains mysterious due to poor quantification of TFP dynamics. Here we fluorescently label the TFP in the opportunistic pathogen Pseudomonas aeruginosa and track the full extension and retraction cycles of individual TFP to quantify their dynamics. We test several models for the switch between extension and retraction using quantitative experiments, biophysical modeling and genetics. We invalidate the prominent hypothesis that this switch is triggered by surface contact. Instead, we show that the entire repetitive cycle of extension and retraction of individual TFP is governed by the stochastic binding of antagonistic extension and retraction motors and explain how this mechanism quantitatively defines physiologically-important features like TFP length and their production rate. Interestingly, our results suggest that the major throttle of TFP production is the unbinding of the retraction motor.

scholarly journals Mechanotaxis directs Pseudomonas aeruginosa twitching motility

2021 ◽  
Author(s):  
Marco J. Kühn ◽  
Lorenzo Talà ◽  
Yuki Inclan ◽  
Ramiro Patino ◽  
Xavier Pierrat ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Single Cells ◽  
Opportunistic Pathogen ◽  
Type Iv Pili ◽  
Twitching Motility ◽  
Response Regulators ◽  
Spatially Resolved ◽  
Type Iv ◽  
Mechanical Signal ◽  
Chp System

AbstractThe opportunistic pathogen Pseudomonas aeruginosa explores surfaces using twitching motility powered by retractile extracellular filaments called type IV pili. Single cells twitch by successive pili extension, attachment and retraction. However, whether and how single cells control twitching migration remains unclear. We discovered that P. aeruginosa actively directs twitching in the direction of mechanical input from type IV pili, in a process we call mechanotaxis. The Chp chemotaxis-like system controls the balance of forward and reverse twitching migration of single cells in response to the mechanical signal. On surfaces, Chp senses type IV pili attachment at one pole thereby sensing a spatially-resolved signal. As a result, the Chp response regulators PilG and PilH control the polarization of the extension motor PilB. PilG stimulates polarization favoring forward migration, while PilH inhibits polarization inducing reversal. Subcellular segregation of PilG and PilH efficiently orchestrates their antagonistic functions, ultimately enabling rapid reversals upon perturbations. This distinct localization of response regulators establishes a signaling landscape known as local-excitation, global-inhibition in higher order organisms, identifying a conserved strategy to transduce spatially-resolved signals. Our discovery finally resolves the function of the Chp system and expands our view of the signals regulating motility.

scholarly journals Competitive binding of independent extension and retraction motors explains the quantitative dynamics of type IV pili

2021 ◽  
Vol 118 (8) ◽  
pp. e2014926118
Author(s):  
Matthias D. Koch ◽  
Chenyi Fei ◽  
Ned S. Wingreen ◽  
Joshua W. Shaevitz ◽  
Zemer Gitai
Keyword(s):  
Production Rate ◽  
Quantitative Agreement ◽  
Type Iv Pili ◽  
Biophysical Model ◽  
Wild Type ◽  
Full Extension ◽  
Multiple Features ◽  
Type Iv ◽  
Quantitative Measurements ◽  
Random Times

Type IV pili (TFP) function through cycles of extension and retraction. The coordination of these cycles remains mysterious due to a lack of quantitative measurements of multiple features of TFP dynamics. Here, we fluorescently label TFP in the pathogen Pseudomonas aeruginosa and track full extension and retraction cycles of individual filaments. Polymerization and depolymerization dynamics are stochastic; TFP are made at random times and extend, pause, and retract for random lengths of time. TFP can also pause for extended periods between two extension or two retraction events in both wild-type cells and a slowly retracting PilT mutant. We developed a biophysical model based on the stochastic binding of two dedicated extension and retraction motors to the same pilus machine that predicts the observed features of the data with no free parameters. We show that only a model in which both motors stochastically bind and unbind to the pilus machine independent of the piliation state of the machine quantitatively explains the experimentally observed pilus production rate. In experimental support of this model, we show that the abundance of the retraction motor dictates the pilus production rate and that PilT is bound to pilus machines even in their unpiliated state. Together, the strong quantitative agreement of our model with a variety of experiments suggests that the entire repetitive cycle of pilus extension and retraction is coordinated by the competition of stochastic motor binding to the pilus machine, and that the retraction motor is the major throttle for pilus production.

scholarly journals Aztreonam Lysine Increases the Activity of Phages E79 and phiKZ against Pseudomonas aeruginosa PA01

2021 ◽  
Vol 9 (1) ◽  
pp. 152
Author(s):  
Carly M. Davis ◽  
Jaclyn G. McCutcheon ◽  
Jonathan J. Dennis
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Morphological Changes ◽  
Burst Size ◽  
Type Iv Pili ◽  
Biofilm Growth ◽  
Promising Alternative ◽  
Type Iv ◽  
Alternative Treatment Option ◽  
One Step ◽  
Step Growth

Pseudomonas aeruginosa is a pernicious bacterial pathogen that is difficult to treat because of high levels of antibiotic resistance. A promising alternative treatment option for such bacteria is the application of bacteriophages; the correct combination of phages plus antibiotics can produce synergistic inhibitory effects. In this study, we describe morphological changes induced by sub-MIC levels of the antibiotic aztreonam lysine (AzLys) on P. aeruginosa PA01, which may in part explain the observed phage–antibiotic synergy (PAS). One-step growth curves for phage E79 showed increased adsorption rates, decreased infection latency, accelerated time to lysis and a minor reduction in burst size. Phage E79 plus AzLys PAS was also able to significantly reduce P. aeruginosa biofilm growth over 3-fold as compared to phage treatment alone. Sub-inhibitory AzLys-induced filamentation of P. aeruginosa cells resulted in loss of twitching motility and a reduction in swimming motility, likely due to a reduction in the number of polar Type IV pili and flagella, respectively, on the filamented cell surfaces. Phage phiKZ, which uses Type IV pili as a receptor, did not exhibit increased activity with AzLys at lower sub-inhibitory levels, but still produced phage–antibiotic synergistic killing with sub-inhibitory AzLys. A one-step growth curve indicates that phiKZ in the presence of AzLys also exhibits a decreased infection latency and moderately undergoes accelerated time to lysis. In contrast to prior PAS studies demonstrating that phages undergo delayed time to lysis with cell filamentation, these PAS results show that phages undergo accelerated time to lysis, which therefore suggests that PAS is dependent upon multiple factors, including the type of phages and antibiotics used, and the bacterial host being tested.

scholarly journals Cryo-ET Characterization of Novel Cellular Extrusions in Escherichia coli Induced by the Major Subunit Protein of Type IV Pili, PilA, from Pseudomonas aeruginosa

2021 ◽  
Vol 27 (S1) ◽  
pp. 280-282
Author(s):  
Juan Sanchez ◽  
Daniel Parrell ◽  
Alba Gonzalez-Rivera ◽  
Nicoleta Ploscariu ◽  
Katrina Forest ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Pseudomonas Aeruginosa ◽  
Type Iv Pili ◽  
Subunit Protein ◽  
Type Iv

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 Disparate Subcellular Localization Patterns of Pseudomonas aeruginosa Type IV Pilus ATPases Involved in Twitching Motility

2005 ◽  
Vol 187 (3) ◽  
pp. 829-839 ◽  
Author(s):  
Poney Chiang ◽  
Marc Habash ◽  
Lori L. Burrows
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Fluorescent Protein ◽  
Protein Localization ◽  
Yellow Fluorescent Protein ◽  
Distribution Patterns ◽  
Opportunistic Pathogen ◽  
Twitching Motility ◽  
Polar Localization ◽  
Type Iv ◽  
And Function

ABSTRACT The opportunistic pathogen Pseudomonas aeruginosa expresses polar type IV pili (TFP), which are responsible for adhesion to various materials and twitching motility on surfaces. Twitching occurs by alternate extension and retraction of TFP, which arise from assembly and disassembly of pilin subunits at the base of the pilus. The ATPase PilB promotes pilin assembly, while the ATPase PilT or PilU or both promote pilin dissociation. Fluorescent fusions to two of the three ATPases (PilT and PilU) were functional, as shown by complementation of the corresponding mutants. PilB and PilT fusions localized to both poles, while PilU fusions localized only to the piliated pole. To identify the portion of the ATPases required for localization, sequential C-terminal deletions of PilT and PilU were generated. The conserved His and Walker B boxes were dispensable for polar localization but were required for twitching motility, showing that localization and function could be uncoupled. Truncated fusions that retained polar localization maintained their distinctive distribution patterns. To dissect the cellular factors involved in establishing polarity, fusion protein localization was monitored with a panel of TFP mutants. The localization of yellow fluorescent protein (YFP)-PilT and YFP-PilU was independent of the subunit PilA, other TFP ATPases, and TFP-associated proteins previously shown to be associated with the membrane or exhibiting polar localization. In contrast, YFP-PilB exhibited diffuse cytoplasmic localization in a pilC mutant, suggesting that PilC is required for polar localization of PilB. Finally, localization studies performed with fluorescent ATPase chimeras of PilT and PilU demonstrated that information responsible for the characteristic localization patterns of the ATPases likely resides in their N termini.

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