scholarly journals Genetic Analysis of the Regulation of Type IV Pilus Function by the Chp Chemosensory System of Pseudomonas aeruginosa

2009 ◽  
Vol 192 (4) ◽  
pp. 994-1010 ◽  
Author(s):  
Jacob J. Bertrand ◽  
Joyce T. West ◽  
Joanne N. Engel
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Histidine Kinase ◽  
Opportunistic Pathogen ◽  
Negative Regulator ◽  
Site Directed Mutagenesis ◽  
Type Iv Pilus ◽  
Chemosensory System ◽  
Type Iv ◽  
Chp System

ABSTRACT The virulence of the opportunistic pathogen Pseudomonas aeruginosa involves the coordinate expression of many virulence factors, including type IV pili, which are required for colonization of host tissues and for twitching motility. Type IV pilus function is controlled in part by the Chp chemosensory system, which includes a histidine kinase, ChpA, and two CheY-like response regulators, PilG and PilH. How the Chp components interface with the type IV pilus motor proteins PilB, PilT, and PilU is unknown. We present genetic evidence confirming the role of ChpA, PilG, and PilB in the regulation of pilus extension and the role of PilH and PilT in regulating pilus retraction. Using informative double and triple mutants, we show that (i) ChpA, PilG, and PilB function upstream of PilH, PilT, and PilU; (ii) that PilH enhances PilT function; and (iii) that PilT and PilB retain some activity in the absence of signaling input from components of the Chp system. By site-directed mutagenesis, we demonstrate that the histidine kinase domain of ChpA and the phosphoacceptor sites of both PilG and PilH are required for type IV pilus function, suggesting that they form a phosphorelay system important in the regulation of pilus extension and retraction. Finally, we present evidence suggesting that pilA transcription is regulated by intracellular PilA levels. We show that PilA is a negative regulator of pilA transcription in P. aeruginosa and that the Chp system functionally regulates pilA transcription by controlling PilA import and export.

scholarly journals The Quorum-Sensing Negative Regulator RsaL of Pseudomonas aeruginosa Binds to the lasI Promoter

2006 ◽  
Vol 188 (2) ◽  
pp. 815-819 ◽  
Author(s):  
Giordano Rampioni ◽  
Iris Bertani ◽  
Elisabetta Zennaro ◽  
Fabio Polticelli ◽  
Vittorio Venturi ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Quorum Sensing ◽  
Binding Site ◽  
Opportunistic Pathogen ◽  
Bidirectional Promoter ◽  
Homoserine Lactone ◽  
Negative Regulator ◽  
Direct Demonstration ◽  
Dna Binding Site

ABSTRACT A mutation in the rsaL gene of Pseudomonas aeruginosa produces dramatically higher amounts of N-acyl homoserine lactone with respect to the wild type, highlighting the key role of this negative regulator in controlling quorum sensing (QS) in this opportunistic pathogen. The DNA binding site of the RsaL protein on the rsaL-lasI bidirectional promoter partially overlaps the binding site of the LasR protein, consistent with the hypothesis that RsaL and LasR could be in binding competition on this promoter. This is the first direct demonstration that RsaL acts as a QS negative regulator by binding to the lasI promoter.

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 ChpC controls twitching motility-mediated expansion of Pseudomonas aeruginosa biofilms in response to serum albumin, mucin and oligopeptides

2019 ◽  
Author(s):  
Laura M. Nolan ◽  
Laura C. McCaughey ◽  
Jessica Merjane ◽  
Lynne Turnbull ◽  
Cynthia B. Whitchurch
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Serum Albumin ◽  
Histidine Kinase ◽  
Cyclic Adenosine Monophosphate ◽  
Adenosine Monophosphate ◽  
Adapter Proteins ◽  
Twitching Motility ◽  
Chemosensory System ◽  
Environmental Signals ◽  
Chp System

AbstractTwitching motility-mediated biofilm expansion occurs via coordinated, multi-cellular collective behaviour to allow bacteria to actively expand across surfaces. Type-IV pili (T4P) are cell-associated virulence factors which mediate this expansion via rounds of extension, surface attachment and retraction. The Chp chemosensory system is thought to respond to environmental signals to regulate the biogenesis, assembly and twitching motility function of T4P. In other well characterised chemosensory systems, methyl-accepting chemotaxis proteins (MCPs) feed environmental signals through a CheW adapter protein to the histidine kinase CheA to modulate motility. The Pseudomonas aeruginosa Chp system has two CheW adapter proteins, PilI and ChpC, and an MCP PilJ that likely interacts via PilI with the histidine kinase ChpA. It is thought that ChpC associates with other MCPs to feed environmental signals into the system, however no such signals have been identified. In the current study we show that ChpC is involved in the response to host-derived signals serum albumin, mucin and oligopeptides. We demonstrate that these signals stimulate an increase in twitching motility, as well as in levels of 3’-5’-cyclic adenosine monophosphate (cAMP) and surface-assembled T4P. Interestingly, our data shows that changes in cAMP and surface piliation levels are independent of ChpC but that the twitching motility response to these environmental signals requires ChpC. Based upon our data we propose a model whereby ChpC associates with an MCP other than PilJ to feed these environmental signals through the Chp system to control twitching motility. The MCP PilJ and the CheW adapter PilI then modulate T4P surface levels to allow the cell to continue to undergo twitching motility. Our study is the first to link environmental signals to the Chp chemosensory system and refines our understanding of how this system controls twitching motility-mediated biofilm expansion in P. aeruginosa.

scholarly journals Functional role of conserved residues in the characteristic secretion NTPase motifs of the Pseudomonas aeruginosa type IV pilus motor proteins PilB, PilT and PilU

Microbiology ◽  
2008 ◽  
Vol 154 (1) ◽  
pp. 114-126 ◽  
Author(s):  
Poney Chiang ◽  
Liliana M. Sampaleanu ◽  
Melissa Ayers ◽  
Markian Pahuta ◽  
P. Lynne Howell ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Functional Role ◽  
Motor Proteins ◽  
Type Iv Pilus ◽  
Conserved Residues ◽  
Type Iv

scholarly journals ChpC controls twitching motility-mediated expansion of Pseudomonas aeruginosa biofilms in response to serum albumin, mucin and oligopeptides

Microbiology ◽  
2020 ◽  
Vol 166 (7) ◽  
pp. 669-678
Author(s):  
Laura M. Nolan ◽  
Laura C. McCaughey ◽  
Jessica Merjane ◽  
Lynne Turnbull ◽  
Cynthia B. Whitchurch
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Serum Albumin ◽  
Histidine Kinase ◽  
Type Species ◽  
Twitching Motility ◽  
Chemosensory System ◽  
Content Type ◽  
Environmental Signals ◽  
Link Type ◽  
Chp System

Twitching motility-mediated biofilm expansion occurs via coordinated, multi-cellular collective behaviour to allow bacteria to actively expand across surfaces. Type-IV pili (T4P) are cell-associated virulence factors which mediate twitching motility via rounds of extension, surface attachment and retraction. The Chp chemosensory system is thought to respond to environmental signals to regulate the biogenesis, assembly and twitching motility function of T4P. In other well characterised chemosensory systems, methyl-accepting chemotaxis proteins (MCPs) feed environmental signals through a CheW adapter protein to the histidine kinase CheA to modulate motility. The Pseudomonas aeruginosa Chp system has an MCP PilJ and two CheW adapter proteins, PilI and ChpC, that likely interact with the histidine kinase ChpA to feed environmental signals into the system. In the current study we show that ChpC is involved in the response to host-derived signals serum albumin, mucin and oligopeptides. We demonstrate that these signals stimulate an increase in twitching motility, as well as in levels of 3′−5′-cyclic adenosine monophosphate (cAMP) and surface-assembled T4P. Interestingly, our data shows that changes in cAMP and surface piliation levels are independent of ChpC but that the twitching motility response to these environmental signals requires ChpC. Furthermore, we show that protease activity is required for the twitching motility response of P. aeruginosa to environmental signals. Based upon our data we propose a model whereby ChpC feeds these environmental signals into the Chp system, potentially via PilJ or another MCP, to control twitching motility. PilJ and PilI then modulate T4P surface levels to allow the cell to continue to undergo twitching motility. Our study is the first to link environmental signals to the Chp chemosensory system and refines our understanding of how this system controls twitching motility-mediated biofilm expansion in P. aeruginosa .

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.

Role of pili in adherence of Pseudomonas aeruginosa to mammalian buccal epithelial cells

1980 ◽  
Vol 29 (3) ◽  
pp. 1146-1151 ◽  
Author(s):  
D E Woods ◽  
D C Straus ◽  
W G Johanson ◽  
V K Berry ◽  
J A Bass
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Epithelial Cells ◽  
Opportunistic Pathogen ◽  
In Vitro System ◽  
Heterologous Antiserum ◽  
Buccal Epithelial Cells ◽  
Serological Studies ◽  
Seriously Ill Patients

Adherence of Pseudomonas aeruginosa organisms to the upper respiratory epithelium of seriously ill patients in vitro is correlated with subsequent colonization of the respiratory tract by this opportunistic pathogen. The role of pili in the attachment to epithelial cells of P. aeruginosa was studied in an in vitro system employing human buccal epithelial cells and P. aeruginosa pretreated by various means. Pretreatment of the bacteria with proteases, heat, or Formalin caused a significant decrease in adherence. A decrease when compared with controls was also noted in the adherence of P. aeruginosa organisms to buccal epithelial cells preincubated with purified pili prepared from the strain used for adherence testing; however, pili prepared from a heterologous strain failed to block adherence. Similar results were obtained in serological studies when antisera to purified pili prepared from the strain used for adherence testing decreased adherence, whereas heterologous antiserum to pili did not decrease adherence. From these results it appears that pili mediate the adherence of P. aeruginosa organisms to human buccal epithelial cells.

scholarly journals The role of core and accessory type IV pilus genes in natural transformation and twitching motility in the bacterium Acinetobacter baylyi

PLoS ONE ◽  
2017 ◽  
Vol 12 (8) ◽  
pp. e0182139 ◽  
Author(s):  
Colleen G. Leong ◽  
Rebecca A. Bloomfield ◽  
Caroline A. Boyd ◽  
Amber J. Dornbusch ◽  
Leah Lieber ◽  
...  
Keyword(s):  
Natural Transformation ◽  
Twitching Motility ◽  
Type Iv Pilus ◽  
Acinetobacter Baylyi ◽  
Type Iv

scholarly journals Pseudomonas aeruginosa-Mediated Damage Requires Distinct Receptors at the Apical and Basolateral Surfaces of the Polarized Epithelium

2009 ◽  
Vol 78 (3) ◽  
pp. 939-953 ◽  
Author(s):  
Iwona Bucior ◽  
Keith Mostov ◽  
Joanne N. Engel
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Mdck Cells ◽  
Three Dimensional ◽  
Opportunistic Pathogen ◽  
Necessary And Sufficient ◽  
Plastic Surfaces ◽  
Pharmacologic Inhibition ◽  
Increased Susceptibility ◽  
Polarized Epithelium

ABSTRACT Pseudomonas aeruginosa, an important opportunistic pathogen of humans, exploits epithelial damage to establish infection. We have rigorously explored the role of N-glycoproteins and heparan sulfate proteoglycans (HSPGs) in P. aeruginosa-mediated attachment and subsequent downstream events at the apical (AP) and basolateral (BL) surfaces of polarized epithelium. We demonstrate that the N-glycan chains at the AP surface are necessary and sufficient for binding, invasion, and cytotoxicity to kidney (MDCK) and airway (Calu-3) cells grown at various states of polarization on Transwell filters. Upregulation of N-glycosylation enhanced binding, whereas pharmacologic inhibition of N-glycosylation or infection of MDCK cells defective in N-glycosylation resulted in decreased binding. In contrast, at the BL surface, the HS moiety of HSPGs mediated P. aeruginosa binding, cytotoxicity, and invasion. In incompletely polarized epithelium, HSPG abundance was increased at the AP surface, explaining its increased susceptibility to P. aeruginosa colonization and damage. Using MDCK cells grown as three-dimensional cysts as a model for epithelial organs, we show that P. aeruginosa specifically colocalized with HS-rich areas at the BL membrane but with complex N-glycans at the AP surface. Finally, P. aeruginosa bound to HS chains and N-glycans coated on plastic surfaces, showing the highest binding affinity toward isolated HS chains. Together, these findings demonstrate that P. aeruginosa recognizes distinct receptors on the AP and BL surfaces of polarized epithelium. Changes in the composition of N-glycan chains and/or in the distribution of HSPGs may explain the enhanced susceptibility of damaged epithelium to P. aeruginosa.

scholarly journals PmtA Regulates Pyocyanin Expression and Biofilm Formation in Pseudomonas aeruginosa

2021 ◽  
Vol 12 ◽  
Author(s):  
Amy V. Thees ◽  
Kathryn M. Pietrosimone ◽  
Clare K. Melchiorre ◽  
Jeremiah N. Marden ◽  
Joerg Graf ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Pseudomonas Aeruginosa ◽  
Metal Binding ◽  
Biofilm Formation ◽  
Binding Capacity ◽  
Opportunistic Pathogen ◽  
Small Molecular Weight ◽  
Heavy Metal Binding ◽  
The Impact

The opportunistic pathogen Pseudomonas aeruginosa expresses a small molecular weight, cysteine-rich protein (PmtA), identified as a metallothionein (MT) protein family member. The MT family proteins have been well-characterized in eukaryotes as essential for zinc and copper homeostasis, protection against oxidative stress, and the ability to modify a variety of immune activities. Bacterial MTs share sequence homology, antioxidant chemistry, and heavy metal-binding capacity with eukaryotic MTs, however, the impact of bacterial MTs on virulence and infection have not been well-studied. In the present study, we investigated the role of PmtA in P. aeruginosa PAO1 using a PmtA-deficient strain (ΔpmtA). Here we demonstrated the virulence factor, pyocyanin, relies on the expression of PmtA. We showed that PmtA may be protective against oxidative stress, as an alternative antioxidant, glutathione, can rescue pyocyanin expression. Furthermore, the expression of phzM, which encodes a pyocyanin precursor enzyme, was decreased in the ΔpmtA mutant during early stationary phase. Upregulated pmtA expression was previously detected in confluent biofilms, which are essential for chronic infection, and we observed that the ΔpmtA mutant was disrupted for biofilm formation. As biofilms also modulate antibiotic susceptibility, we examined the ΔpmtA mutant susceptibility to antibiotics and found that the ΔpmtA mutant is more susceptible to cefepime and ciprofloxacin than the wild-type strain. Finally, we observed that the deletion of pmtA results in decreased virulence in a waxworm model. Taken together, our results support the conclusion that PmtA is necessary for the full virulence of P. aeruginosa and may represent a potential target for therapeutic intervention.

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