scholarly journals Obstruction of pilus retraction stimulates bacterial surface sensing

2017 ◽  
Author(s):  
Courtney K. Ellison ◽  
Jingbo Kan ◽  
Rebecca S. Dillard ◽  
David T. Kysela ◽  
Cheri M. Hampton ◽  
...  
Keyword(s):  
Genetic Modification ◽  
Caulobacter Crescentus ◽  
Physiological Changes ◽  
Type Iv Pilus ◽  
Dynamic Nature ◽  
Surface Attachment ◽  
Bacterial Surface ◽  
Type Iv ◽  
Surface Sensing ◽  
Surface Contact

AbstractSurface association provides numerous fitness advantages to bacteria. Thus, it is critical for bacteria to recognize surface contact and to consequently initiate physiological changes required for a surface-associated lifestyle (1). Ubiquitous microbial appendages called pili are involved in sensing surfaces and mediating downstream surface-associated behaviors (2–6). The mechanism by which pili mediate surface sensing remains unknown, largely due to the difficulty to visualize their dynamic nature and to directly modulate their activity without genetic modification. Here, we show thatCaulobacter crescentuspili undergo dynamic cycles of extension and retraction that cease within seconds of surface contact, and this arrest of pilus activity coincides with surface-stimulated holdfast synthesis. By physically blocking pili, we show that imposing resistance to pilus retraction is sufficient to stimulate holdfast synthesis in the absence of surface contact. Thus, resistance to type IV pilus retraction upon surface attachment is used for surface sensing.One Sentence SummaryBacteria use the tension imparted on retracting pilus fibers upon their binding to a surface for surface sensing.

scholarly journals More than a feeling: microscopy approaches to understanding surface-sensing mechanisms

2020 ◽  
Author(s):  
Katherine J. Graham ◽  
Lori L. Burrows
Keyword(s):  
Live Cells ◽  
Surface Attachment ◽  
Bacterial Surface ◽  
Structural Modifications ◽  
Structure And Dynamics ◽  
Type Iv ◽  
Surface Sensing ◽  
Two Component Signal Transduction ◽  
Sense And Respond

The mechanisms by which bacteria sense and respond to surface attachment have long been a mystery. Our understanding of the structure and dynamics of bacterial appendages, notably type IV pili (T4P), provided new insights into the potential ways that bacteria sense surfaces. T4P are ubiquitous, retractable hair-like adhesins that until recently were difficult to image in the absence of fixation due to their nanoscale size. This review focuses on recent microscopy innovations used to visualize T4P in live cells to reveal the dynamics of their retraction and extension. We discuss recently proposed mechanisms by which T4P facilitate bacterial surface sensing, including the role of surface-exposed PilY1, two-component signal transduction pathways, force-induced structural modifications of the major pilin, and altered dynamics of the T4P motor complex.

scholarly journals The type IV pilin PilA couples surface attachment and cell cycle initiation in Caulobacter crescentus

2019 ◽  
Author(s):  
Luca Del Medico ◽  
Dario Cerletti ◽  
Matthias Christen ◽  
Beat Christen
Keyword(s):  
Cell Cycle ◽  
Biofilm Formation ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Peptide Mimetics ◽  
Cycle Progression ◽  
Surface Attachment ◽  
Type Iv ◽  
Surface Sensing ◽  
Type Iv Pilin

Understanding how bacteria colonize surfaces and regulate cell cycle progression in response to cellular adhesion is of fundamental importance. Here, we used transposon sequencing in conjunction with FRET microscopy to uncover the molecular mechanism how surface sensing drives cell cycle initiation in Caulobacter crescentus. We identified the type IV pilin protein PilA as the primary signaling input that couples surface contact to cell cycle initiation via the second messenger c-di-GMP. Upon retraction of pili filaments, the monomeric pilin reservoir in the inner membrane is sensed by the 17 amino-acid transmembrane helix of PilA to activate the PleC-PleD two component signaling system, increase cellular c-di-GMP levels and signal the onset of the cell cycle. We termed the PilA signaling sequence CIP for cell cycle initiating pilin peptide. Addition of the chemically synthesized CIP peptide initiates cell cycle progression and simultaneously inhibits surface attachment. The broad conservation of the type IV pili and their importance in pathogens for host colonization suggests that CIP peptide mimetics offer new strategies to inhibit surface-sensing, prevent biofilm formation and control persistent infections.Significance StatementPili are hair-like appendages found on the surface of many bacteria to promote adhesion. Here, we provide systems-level findings on a molecular signal transduction pathway that interlinks surface sensing with cell cycle initiation. We propose that surface attachment induces depolymerization of pili filaments. The concomitant increase in pilin sub-units within the inner membrane function as a stimulus to activate the second messenger c-di-GMP and trigger cell cycle initiation. Further-more, we show that the provision of a 17 amino acid synthetic peptide corresponding to the membrane portion of the pilin sub-unit mimics surface sensing, activates cell cycle initiation and inhibits surface attachment. Thus, synthetic peptide mimetics of pilin may represent new chemotypes to control biofilm formation and treat bacterial infections.

scholarly journals A bifunctional ATPase drives tad pilus extension and retraction

2019 ◽  
Author(s):  
Courtney K. Ellison ◽  
Jingbo Kan ◽  
Jennifer L. Chlebek ◽  
Katherine R. Hummels ◽  
Gaёl Panis ◽  
...  
Keyword(s):  
Molecular Motors ◽  
Intracellular Transport ◽  
Atp Hydrolysis ◽  
Caulobacter Crescentus ◽  
Dynamic Activity ◽  
Surface Attachment ◽  
Type Iv ◽  
Surface Sensing ◽  
Domains Of Life ◽  
Macromolecular Protein

AbstractMolecular motors convert chemical energy directly into mechanical work1and are found in all domains of life2. These motors are critical to intracellular transport3, motility4,5, macromolecular protein assembly3,6, and many essential processes7. A wide-spread class of related bacterial motors drive the dynamic activity of extracellular fibers, such as type IV pili (T4P), that are extended and retracted using so-called secretion motor ATPases. Among these, the tightadherence (tad) pili are critical for surface sensing, surface attachment, and biofilm formation8–10. How tad pili undergo dynamic cycles of extension and retraction8despite lacking a dedicated retraction motor ATPase has remained a mystery. Here we find that a bifunctional pilus motor ATPase, CpaF, drives both activities through ATP hydrolysis. Specifically, we show that mutations within the ATP hydrolysis active site ofCaulobacter crescentusCpaF result in a correlated reduction in the rates of extension and retraction. Moreover, a decrease in the rate of ATP hydrolysis directly scales with a decrease in the force of retraction and reduced dynamics in these CpaF mutants. This mechanism of motor protein bifunctionality extends to another genus of tad-bearing bacteria. In contrast, the T4aP subclass of pili possess dedicated extension and retraction motor ATPase paralogs. We show that these processes are uncoupled using a slow ATP hydrolysis mutation in the extension ATPase of competence T4aP ofVibrio choleraethat decreases the rate of extension but has no effect on the rate of retraction. Thus, a single motor ATPase is able to drive the bidirectional processes of pilus fiber extension and retraction.

scholarly journals The type IV pilin PilA couples surface attachment and cell-cycle initiation in Caulobacter crescentus

2020 ◽  
Vol 117 (17) ◽  
pp. 9546-9553 ◽  
Author(s):  
Luca Del Medico ◽  
Dario Cerletti ◽  
Philipp Schächle ◽  
Matthias Christen ◽  
Beat Christen
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Resonance Energy ◽  
Cellular Adhesion ◽  
Cycle Progression ◽  
Surface Attachment ◽  
Type Iv ◽  
Surface Sensing ◽  
Type Iv Pilin

Understanding how bacteria colonize surfaces and regulate cell-cycle progression in response to cellular adhesion is of fundamental importance. Here, we use transposon sequencing in conjunction with fluorescence resonance energy transfer (FRET) microscopy to uncover the molecular mechanism for how surface sensing drives cell-cycle initiation in Caulobacter crescentus. We identify the type IV pilin protein PilA as the primary signaling input that couples surface contact to cell-cycle initiation via the second messenger cyclic di-GMP (c-di-GMP). Upon retraction of pili filaments, the monomeric pilin reservoir in the inner membrane is sensed by the 17-amino acid transmembrane helix of PilA to activate the PleC-PleD two-component signaling system, increase cellular c-di-GMP levels, and signal the onset of the cell cycle. We termed the PilA signaling sequence CIP for “cell-cycle initiating pilin” peptide. Addition of the chemically synthesized CIP peptide initiates cell-cycle progression and simultaneously inhibits surface attachment. The broad conservation of the type IV pili and their importance in pathogens for host colonization suggests that CIP peptide mimetics offer strategies to inhibit surface sensing, prevent biofilm formation and control persistent infections.

The Xanthomonas oryzae pv. oryzae type IV pilus alignment subcomplex protein PilN contributes to regulation of bacterial surface‐associated behaviours and T3SS system

2020 ◽  
Vol 69 (4) ◽  
pp. 744-755
Author(s):  
Yilang Li ◽  
Yichao Yan ◽  
Songge Deng ◽  
Cuiping Zhang ◽  
Fazal Haq ◽  
...  
Keyword(s):  
Xanthomonas Oryzae ◽  
Type Iv Pilus ◽  
Bacterial Surface ◽  
Type Iv

scholarly journals Surface Sensing Stimulates Cellular Differentiation in Caulobacter crescentus

2019 ◽  
Author(s):  
Rhett A. Snyder ◽  
Courtney K. Ellison ◽  
Geoffrey B. Severin ◽  
Christopher M. Waters ◽  
Yves V. Brun
Keyword(s):  
Cell Cycle ◽  
Cell Differentiation ◽  
Cellular Differentiation ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Chromosome Replication ◽  
Fundamental Aspect ◽  
Cycle Progression ◽  
Surface Sensing ◽  
Surface Contact

AbstractCellular differentiation is a fundamental strategy used by cells to generate specialized functions at specific stages of development. The bacterium C. crescentus employs a specialized dimorphic life cycle consisting of two differentiated cell types. How environmental cues, including mechanical inputs such as contact with a surface, regulate this cell cycle remain unclear. Here, we find that surface sensing by the physical perturbation of retracting extracellular pilus filaments accelerates cell cycle progression and cellular differentiation. We show that physical obstruction of dynamic pilus activity by chemical perturbation or by a mutation in the outer membrane pilus pore protein, CpaC, stimulates early initiation of chromosome replication. In addition, we find that surface contact stimulates cell cycle progression by demonstrating that surface-stimulated cells initiate early chromosome replication to the same extent as planktonic cells with obstructed pilus activity. Finally, we show that obstruction of pilus retraction stimulates the synthesis of the cell cycle regulator, cyclic diguanylate monophosphate (c-di-GMP) through changes in the activity and localization of two key regulatory histidine kinases that control cell fate and differentiation. Together, these results demonstrate that surface contact and mechanosensing by alterations in pilus activity stimulate C. crescentus to bypass its developmentally programmed temporal delay in cell differentiation to more quickly adapt to a surface-associated lifestyle.SignificanceCells from all domains of life sense and respond to mechanical cues [1–3]. In eukaryotes, mechanical signals such as adhesion and surface stiffness are important for regulating fundamental processes including cell differentiation during embryonic development [4]. While mechanobiology is abundantly studied in eukaryotes, the role of mechanical influences on prokaryotic biology remains under-investigated. Here, we demonstrate that mechanosensing mediated through obstruction of the dynamic extension and retraction of tight adherence (tad) pili stimulates cell differentiation and cell cycle progression in the dimorphic α-proteobacterium Caulobacter crescentus. Our results demonstrate an important intersection between mechanical stimuli and the regulation of a fundamental aspect of cell biology.

scholarly journals Tad pili play a dynamic role in Caulobacter crescentus surface colonization

2019 ◽  
Author(s):  
Matteo Sangermani ◽  
Isabelle Hug ◽  
Nora Sauter ◽  
Thomas Pfohl ◽  
Urs Jenal
Keyword(s):  
Caulobacter Crescentus ◽  
Optical Trap ◽  
Natural Environments ◽  
Surface Attachment ◽  
Surface Colonization ◽  
Bacterial Surface ◽  
Controlled Flow ◽  
Mechanical Sensing

ABSTRACTBacterial surface attachment is mediated by rotary flagella and filamentous appendages called pili. Here, we describe the role of Tad pili during surface colonization of Caulobacter crescentus. Using an optical trap and microfluidic controlled flow conditions as a mimic of natural environments, we demonstrate that Tad pili undergo repeated cycles of extension and retraction. Within seconds after establishing surface contact, pili reorient cells into an upright position promoting walking-like movements against the medium flow. Pili-mediated positioning of the flagellated pole close to the surface facilitates motor-mediated mechanical sensing and promotes anchoring of the holdfast, an adhesive substance that affords long-term attachment. We present evidence that the second messenger c-di-GMP regulates pili dynamics during surface encounter in distinct ways, promoting increased activity at intermediate levels and retraction of pili at peak concentrations. We propose a model, in which flagellum and Tad pili functionally interact and together impose a ratchet-like mechanism that progressively drives C. crescentus cells towards permanent surface attachment.

scholarly journals Social Cooperativity of Bacteria during Reversible Surface Attachment in Young Biofilms: a Quantitative Comparison of Pseudomonas aeruginosa PA14 and PAO1

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Calvin K. Lee ◽  
Jérémy Vachier ◽  
Jaime de Anda ◽  
Kun Zhao ◽  
Amy E. Baker ◽  
...  
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Stochastic Model ◽  
Bacterial Biofilm ◽  
Population Increase ◽  
Pivotal Phase ◽  
Surface Attachment ◽  
Content Type ◽  
Cellular Behavior ◽  
Bacterial Surface ◽  
Surface Sensing

ABSTRACT What are bacteria doing during “reversible attachment,” the period of transient surface attachment when they initially engage a surface, besides attaching themselves to the surface? Can an attaching cell help any other cell attach? If so, does it help all cells or employ a more selective strategy to help either nearby cells (spatial neighbors) or its progeny (temporal neighbors)? Using community tracking methods at the single-cell resolution, we suggest answers to these questions based on how reversible attachment progresses during surface sensing for Pseudomonas aeruginosa strains PAO1 and PA14. Although PAO1 and PA14 exhibit similar trends of surface cell population increase, they show unanticipated differences when cells are considered at the lineage level and interpreted using the quantitative framework of an exactly solvable stochastic model. Reversible attachment comprises two regimes of behavior, processive and nonprocessive, corresponding to whether cells of the lineage stay on the surface long enough to divide, or not, before detaching. Stark differences between PAO1 and PA14 in the processive regime of reversible attachment suggest the existence of two surface colonization strategies. PAO1 lineages commit quickly to a surface compared to PA14 lineages, with early c-di-GMP-mediated exopolysaccharide (EPS) production that can facilitate the attachment of neighbors. PA14 lineages modulate their motility via cyclic AMP (cAMP) and retain memory of the surface so that their progeny are primed for improved subsequent surface attachment. Based on the findings of previous studies, we propose that the differences between PAO1 and PA14 are potentially rooted in downstream differences between Wsp-based and Pil-Chp-based surface-sensing systems, respectively. IMPORTANCE The initial pivotal phase of bacterial biofilm formation known as reversible attachment, where cells undergo a period of transient surface attachment, is at once universal and poorly understood. What is more, although we know that reversible attachment culminates ultimately in irreversible attachment, it is not clear how reversible attachment progresses phenotypically, as bacterial surface-sensing circuits fundamentally alter cellular behavior. We analyze diverse observed bacterial behavior one family at a time (defined as a full lineage of cells related to one another by division) using a unifying stochastic model and show that our findings lead to insights on the time evolution of reversible attachment and the social cooperative dimension of surface attachment in PAO1 and PA14 strains.

scholarly journals Surface sensing stimulates cellular differentiation inCaulobacter crescentus

2020 ◽  
Vol 117 (30) ◽  
pp. 17984-17991 ◽  
Author(s):  
Rhett A. Snyder ◽  
Courtney K. Ellison ◽  
Geoffrey B. Severin ◽  
Gregory B. Whitfield ◽  
Christopher M. Waters ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cell Fate ◽  
Cellular Differentiation ◽  
Cell Cycle Progression ◽  
Caulobacter Crescentus ◽  
Control Cell ◽  
Chromosome Replication ◽  
Cycle Progression ◽  
Surface Sensing ◽  
Surface Contact

Cellular differentiation is a fundamental strategy used by cells to generate specialized functions at specific stages of development. The bacteriumCaulobacter crescentusemploys a specialized dimorphic life cycle consisting of two differentiated cell types. How environmental cues, including mechanical inputs such as contact with a surface, regulate this cell cycle remain unclear. Here, we find that surface sensing by the physical perturbation of retracting extracellular pilus filaments accelerates cell-cycle progression and cellular differentiation. We show that physical obstruction of dynamic pilus activity by chemical perturbation or by a mutation in the outer-membrane pilus secretin CpaC stimulates early initiation of chromosome replication. In addition, we find that surface contact stimulates cell-cycle progression by demonstrating that surface-stimulated cells initiate early chromosome replication to the same extent as planktonic cells with obstructed pilus activity. Finally, we show that obstruction of pilus retraction stimulates the synthesis of the cell-cycle regulator cyclic diguanylate monophosphate (c-di-GMP) through changes in the activity and localization of two key regulatory histidine kinases that control cell fate and differentiation. Together, these results demonstrate that surface contact and sensing by alterations in pilus activity stimulateC. crescentusto bypass its developmentally programmed temporal delay in cell differentiation to more quickly adapt to a surface-associated lifestyle.

scholarly journals Cyclic di-GMP differentially tunes a bacterial flagellar motor through a novel class of CheY-like regulators

eLife ◽  
2017 ◽  
Vol 6 ◽  
Author(s):  
Jutta Nesper ◽  
Isabelle Hug ◽  
Setsu Kato ◽  
Chee-Seng Hee ◽  
Judith Maria Habazettl ◽  
...  
Keyword(s):  
Motor Activity ◽  
Caulobacter Crescentus ◽  
Chemical Information ◽  
Flagellar Motor ◽  
Cellular Functions ◽  
Surface Attachment ◽  
Chemical Proteomics ◽  
Bacterial Surface ◽  
Rotary Machine ◽  
Motile Cells

The flagellar motor is a sophisticated rotary machine facilitating locomotion and signal transduction. Owing to its important role in bacterial behavior, its assembly and activity are tightly regulated. For example, chemotaxis relies on a sensory pathway coupling chemical information to rotational bias of the motor through phosphorylation of the motor switch protein CheY. Using a chemical proteomics approach, we identified a novel family of CheY-like (Cle) proteins in Caulobacter crescentus, which tune flagellar activity in response to binding of the second messenger c-di-GMP to a C-terminal extension. In their c-di-GMP bound conformation Cle proteins interact with the flagellar switch to control motor activity. We show that individual Cle proteins have adopted discrete cellular functions by interfering with chemotaxis and by promoting rapid surface attachment of motile cells. This study broadens the regulatory versatility of bacterial motors and unfolds mechanisms that tie motor activity to mechanical cues and bacterial surface adaptation.

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