PilJ Localizes to Cell Poles and Is Required for Type IV Pilus Extension in Pseudomonas aeruginosa

DeLange, Paul A.; Collins, Tracy; Pierce, George E.; Robinson, Jayne B.

doi:10.1007/s00284-007-9008-5

Cited by 33 publications

(29 citation statements)

References 28 publications

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“…Targets for future study could include pilus extension or retraction motor proteins, e.g. ATPases PilB or PilT respectively [49, 50], or the chemosensory protein PilJ which directly interacts with PilA [23], and also controls pilus extension [51]. …”

Section: Discussionmentioning

confidence: 99%

Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

Metruccio

Evans

et al. 2017

PLoS Pathog

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It is generally thought that mucosal fluids protect underlying epithelial surfaces against opportunistic infection via their antimicrobial activity. However, our published data show that human tear fluid can protect against the major opportunistic pathogen Pseudomonas aeruginosa independently of bacteriostatic activity. Here, we explored the mechanisms for tear protection, focusing on impacts of tear fluid on bacterial virulence factor expression. Results showed that tear fluid suppressed twitching motility, a type of surface-associated movement conferred by pili. Previously, we showed that twitching is critical for P. aeruginosa traversal of corneal epithelia, exit from epithelial cells after internalization, and corneal virulence. Inhibition of twitching by tear fluid was dose-dependent with dilutions to 6.25% retaining activity. Purified lactoferrin, lysozyme, and contrived tears containing these, and many other, tear components lacked the activity. Systematic protein fractionation, mass spectrometry, and immunoprecipitation identified the glycoprotein DMBT1 (Deleted in Malignant Brain Tumors 1) in tear fluid as required. DMBT1 purified from human saliva also inhibited twitching, as well as P. aeruginosa traversal of human corneal epithelial cells in vitro, and reduced disease pathology in a murine model of corneal infection. DMBT1 did not affect PilA expression, nor bacterial intracellular cyclicAMP levels, and suppressed twitching motility of P. aeruginosa chemotaxis mutants (chpB, pilK), and an adenylate cyclase mutant (cyaB). However, dot-immunoblot assays showed purified DMBT1 binding of pili extracted from PAO1 suggesting that twitching inhibition may involve a direct interaction with pili. The latter could affect extension or retraction of pili, their interactions with biotic or abiotic surfaces, or cause their aggregation. Together, the data suggest that DMBT1 inhibition of twitching motility contributes to the mechanisms by which mucosal fluids protect against P. aeruginosa infection. This study also advances our understanding of how mucosal fluids protect against infection, and suggests directions for novel biocompatible strategies to protect our surface epithelia against a major opportunistic pathogen.

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Section: Discussionmentioning

confidence: 99%

Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

Metruccio

Evans

et al. 2017

PLoS Pathog

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“…CyaB is localized in the bacterial poles 40 , a location shared by the tfp biogenesis machinery 41 and components of the Chp chemosensory system 40; 42 . We predict that the MASE2 domain, which we show here is required for inner membrane targeting, is also likely to be required for localization to the bacterial poles.…”

Section: Discussionmentioning

confidence: 99%

Crystal Structure and Regulation Mechanisms of the CyaB Adenylyl Cyclase from the Human Pathogen Pseudomonas aeruginosa

Topal

Fulcher

Bitterman

et al. 2012

Journal of Molecular Biology

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Pseudomonas aeruginosa is an opportunistic bacterial pathogen and major cause of healthcare-associated infections. While the organism’s intrinsic and acquired resistance to most antibiotics hinders treatment of P. aeruginosa infections, the regulatory networks controlling its virulence provide novel targets for drug development. CyaB, a key regulator of P. aeruginosa virulence, belongs to the Class III adenylyl cyclase (AC) family of enzymes that synthesize the second messenger cyclic adenosine 3′,5′-monophosphate (cAMP). These enzymes consist of a conserved catalytic domain fused to one or more regulatory domains. We describe here the biochemical and structural characterization of CyaB and its inhibition by small molecules. We show that CyaB belongs to the Class IIIb subfamily and, like other subfamily members, its activity is stimulated by inorganic carbon. CyaB is also regulated by its N-terminal MASE2 domain, which acts as a membrane anchor. Using a genetic screen, we identified activating mutations in CyaB. By solving the crystal structure of the CyaB catalytic domain, we rationalized the effects of these mutations and propose that CyaB employs regulatory mechanisms similar to other Class III ACs. The CyaB structure further indicates subtle differences compared to other Class III ACs in both the active and inhibitor binding pocket. Consistent with these differences, we observed a unique inhibition profile, including identification of a CyaB selective compound. Overall, our results reveal mechanistic details of the physiological and pharmacological regulation of CyaB and provide the basis for its exploitation as a therapeutic drug target.

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“…Attached TFP are under tension during retraction and this tension could modify TFP, for example by inducing a modification in PilA. PilJ could sense such tension-induced changes at the attachment point between the cell body and the TFP, where PilJ colocalizes (25). Alternatively, PilJ could sense TFP modification during retraction, as PilA subunits depolymerize into monomers and are incorporated back into the inner membrane.…”

Section: Mechanotransduction Requires Both Tfp Tension and Retractionmentioning

confidence: 99%

Type IV pili mechanochemically regulate virulence factors in Pseudomonas aeruginosa

Persat

Inclán

Engel

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

353

398

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Bacteria have evolved a wide range of sensing systems to appropriately respond to environmental signals. Here we demonstrate that the opportunistic pathogen Pseudomonas aeruginosa detects contact with surfaces on short timescales using the mechanical activity of its type IV pili, a major surface adhesin. This signal transduction mechanism requires attachment of type IV pili to a solid surface, followed by pilus retraction and signal transduction through the Chp chemosensory system, a chemotaxis-like sensory system that regulates cAMP production and transcription of hundreds of genes, including key virulence factors. Like other chemotaxis pathways, pili-mediated surface sensing results in a transient response amplified by a positive feedback that increases type IV pili activity, thereby promoting long-term surface attachment that can stimulate additional virulence and biofilm-inducing pathways. The methyl-accepting chemotaxis protein-like chemosensor PilJ directly interacts with the major pilin subunit PilA. Our results thus support a mechanochemical model where a chemosensory system measures the mechanically induced conformational changes in stretched type IV pili. These findings demonstrate that P. aeruginosa not only uses type IV pili for surface-specific twitching motility, but also as a sensor regulating surface-induced gene expression and pathogenicity.surface sensing | mechanotransduction | type IV pili | virulence N umerous bacterial species exhibit a range of behaviors that are specific to life on surfaces. For many pathogens, surfacespecific phenotypes are often associated with virulent activity, because many infection strategies require contact with a host (1). For example, in Pseudomonas aeruginosa, the type III secretion system, which infects by injecting toxins, requires attachment of individual cells to host-cell membranes (2). The genetic pathways that regulate P. aeruginosa secretion systems have been well characterized (3), but the environmental signals that activate these pathways remain poorly defined (4). Given the requirement for host-cell contact for efficient infection, P. aeruginosa could leverage surface contact as a signal to properly activate and coordinate pathogenicity. Indeed, our laboratory recently demonstrated that prolonged association with a solid surface contributes to P. aeruginosa pathogenicity when bacteria and host cells are forced together (5). Here we address the fundamental question of how bacteria rapidly sense surface contact and transduce this input into a cellular response over short timescales.Surface-specific behaviors require intimate contact between cells and substrate. In P. aeruginosa, contact is mediated by several adhesins, particularly type IV pili (TFP). TFP are long, motorized fimbriae that also provide cells with surface-specific twitching motility and are essential to virulence and biofilm formation (6, 7). Successive TFP extension, attachment, and retraction promote intimate association with surfaces and motility along them. Because TFP dynamically int...

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PilJ Localizes to Cell Poles and Is Required for Type IV Pilus Extension in Pseudomonas aeruginosa

Cited by 33 publications

References 28 publications

Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

Mucosal fluid glycoprotein DMBT1 suppresses twitching motility and virulence of the opportunistic pathogen Pseudomonas aeruginosa

Crystal Structure and Regulation Mechanisms of the CyaB Adenylyl Cyclase from the Human Pathogen Pseudomonas aeruginosa

Type IV pili mechanochemically regulate virulence factors in Pseudomonas aeruginosa

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