Layered Structure and Complex Mechanochemistry Underlie Strength and Versatility in a Bacterial Adhesive

Hernando-Pérez, Mercedes; Setayeshgar, Sima; Hou, Yifeng; Témam, Roger; Brun, Yves V.; Dragnea, Bogdan; Berne, Christian

doi:10.1128/mbio.02359-17

Cited by 29 publications

(56 citation statements)

References 87 publications

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“…S2). It has been shown recently that the amine-reactive succinimidyl ester dye shows the same labeling profile (Hernando-Pérez et al, 2018). Shed holdfasts from a ∆hfaB mutant differ from holdfasts anchored to WT cells in that they lack the cysteine-containing outer membrane holdfast anchor protein HfaD (Hardy et al, 2010).…”

Section: Surface Contact Stimulation Of Holdfast Synthesis Is Not Impmentioning

confidence: 99%

Feedback regulation of Caulobacter crescentus holdfast synthesis by flagellum assembly via the holdfast inhibitor HfiA

Berne

Ellison

Agarwal

et al. 2018

Molecular Microbiology

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To permanently attach to surfaces, Caulobacter crescentusproduces a strong adhesive, the holdfast. The timing of holdfast synthesis is developmentally regulated by cell cycle cues. When C. crescentusis grown in a complex medium, holdfast synthesis can also be stimulated by surface sensing, in which swarmer cells rapidly synthesize holdfast in direct response to surface contact. In contrast to growth in complex medium, here we show that when cells are grown in a defined medium, surface contact does not trigger holdfast synthesis. Moreover, we show that in a defined medium, flagellum synthesis and regulation of holdfast production are linked. In these conditions, mutants lacking a flagellum attach to surfaces over time more efficiently than either wild-type strains or strains harboring a paralyzed flagellum. Enhanced adhesion in mutants lacking flagellar components is due to premature holdfast synthesis during the cell cycle and is regulated by the holdfast synthesis inhibitor HfiA. hfiA transcription is reduced in flagellar mutants and this reduction is modulated by the diguanylate cyclase developmental regulator PleD. We also show that, in contrast to previous predictions, flagella are not necessarily required for C. crescentus surface sensing in the absence of flow, and that arrest of flagellar rotation does not stimulate holdfast synthesis. Rather, our data support a model in which flagellum assembly feeds back to control holdfast synthesis via HfiA expression in a c-di-GMP-dependent manner under defined nutrient conditions.

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Section: Surface Contact Stimulation Of Holdfast Synthesis Is Not Impmentioning

confidence: 99%

Feedback regulation of Caulobacter crescentus holdfast synthesis by flagellum assembly via the holdfast inhibitor HfiA

Berne

Ellison

Agarwal

et al. 2018

Molecular Microbiology

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“…the holdfast, but only mutations in polysaccharide biosynthesis genes or pleiotropic regulators of cell polarity abolish holdfast production (24). Currently, three glycosyltransferase steps are known to be required for holdfast synthesis.…”

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confidence: 99%

A genome-wide analysis of adhesion inCaulobacter crescentusidentifies new regulatory and biosynthetic components for holdfast assembly

Hershey

Fiebig

Crosson

2018

Preprint

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Due to their intimate physical interactions with the environment, surface polysaccharides are critical determinants of fitness for bacteria. Caulobacter crescentus produces a specialized structure at one of its cell poles called the holdfast that enables attachment to surfaces. Previous studies have shown that the holdfast is a carbohydrate-based material and identified a number of genes required for holdfast development. However, incomplete information about its chemical structure, biosynthetic genes and regulatory principles has limited progress in understanding the mechanism of holdfast synthesis. We have leveraged the adhesive properties of the holdfast to perform a saturating screen for genes affecting attachment to cheesecloth over a multi-day time course. Using similarities in the temporal profiles of mutants in a transposon library, we defined discrete clusters of genes with related effects on cheesecloth colonization. Holdfast synthesis, flagellar motility, type IV pilus assembly and smooth lipopolysaccharide (SLPS) production represented key classes of adhesion determinants. Examining these clusters in detail allowed us to predict and experimentally define the functions of multiple uncharacterized genes in both the holdfast and SLPS pathways. In addition, we showed that the pilus and flagellum control holdfast synthesis separately by modulating the holdfast inhibitor hfiA. This study defines a set of genes contributing to adhesion that includes newly discovered genes required for holdfast biosynthesis and attachment. Our data provide evidence that the holdfast contains a complex polysaccharide with at least four monosaccharides in the repeating unit and underscore the central role of cell polarity in mediating attachment of C. crescentus to surfaces. Importance: Bacteria routinely encounter biotic and abiotic materials in their surrounding environments, and they often enlist specific behavioral programs to colonize these materials. Adhesion is an early step in colonizing a surface. Caulobacter crescentus produces a structure called the holdfast, which allows this organism to attach to and colonize surfaces. To understand how the holdfast is produced, we performed a genome-wide search for genes that contribute to adhesion by selecting for mutants that could not attach to cheesecloth. We discovered complex interactions between genes that mediate surface contact and genes that contribute to holdfast development. Our genetic selection identified what likely represents a comprehensive set of genes required to generate a holdfast, laying the groundwork for a detailed characterization of the enzymes that build this specialized adhesin.

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“…This molecule is a wellestablished component of the biofilm matrix of other bacteria (17). DNA associates with the outer layers of the C. crescentus holdfast and similarly is observed adjacent to the holdfast polysaccharide in rosette cores (33). In other work, DNA released during cell death was demonstrated to bind to holdfast and inhibit attachment (67).…”

Section: On the Formation Of Strings Of Cells And The Threads That Comentioning

confidence: 92%

“…Lectin staining and enzymatic digestion studies indicate Nacetylglucosamine moieties are present in the holdfast (32), and there is also evidence that protein and DNA are important components of this adhesin (33). The role of the C. crescentus 3 holdfast and other surface structures, including the flagellum and type IV pili, in colonization of the air-liquid interface has not been investigated.…”

Section: A Dimorphic Bacterial Model To Study Colonization Of the Airmentioning

confidence: 99%

The role ofCaulobactercell surface structures in colonization of the air-liquid interface

Fiebig

2019

Preprint

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In aquatic environments, Caulobacter spp. are often present at the boundary between liquid and air known as the neuston. I report an approach to study temporal features of Caulobacter crescentus colonization and pellicle biofilm development at the air-liquid interface, and have defined the role of cell surface structures in this process. The flagellum enables motile swarmer cells to efficiently reach the oxygenated surface. Here, cells form a monolayer enriched in stalked cells bearing a surface adhesin known as a holdfast. When excised from the liquid surface, this monolayer strongly adheres to glass. The monolayer subsequently develops into a three-dimensional structure that is highly enriched in clusters of stalked cells known as rosettes. As the pellicle film matures, it becomes more cohesive and less adherent to a glass surface. A mutant strain lacking a flagellum does not efficiently reach the surface, and strains lacking type IV pili exhibit defects in organization of the three-dimensional pellicle. Strains unable to synthesize holdfast fail to accumulate at the air-liquid interface and do not form a pellicle. Phase contrast images support a model whereby the holdfast functions to trap C. crescentus cells at the air-liquid boundary. Unlike the holdfast, neither the flagellum nor pili are required for C. crescentus to partition to the air-liquid interface. While it is well established that the holdfast enables adherence to solid surfaces, this study provides evidence that the holdfast has physicochemical properties that enable partitioning of non-motile mother cells to the air-liquid interface, which facilitates colonization of this microenvironment. ImportanceIn aquatic environments the boundary at the air interface is often highly enriched with nutrients and oxygen. The ability of microbial cells to colonize this niche likely confers a significant fitness advantage in many cases. This study provides evidence that the cell surface adhesin known as a holdfast enables Caulobacter crescentus to partition to and colonize the air-liquid interface. Additional surface structures including the flagellum and pili are important determinants of colonization and biofilm formation at this boundary. Considering that holdfast-like adhesins are broadly conserved in Caulobacter spp. and other members of the diverse class Alphaproteobacteria, these surface structures may function broadly to facilitate colonization of air-liquid boundaries in a range of ecological contexts including freshwater, marine, and soil ecosystems.

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Layered Structure and Complex Mechanochemistry Underlie Strength and Versatility in a Bacterial Adhesive

Cited by 29 publications

References 87 publications

Feedback regulation of Caulobacter crescentus holdfast synthesis by flagellum assembly via the holdfast inhibitor HfiA

Feedback regulation of Caulobacter crescentus holdfast synthesis by flagellum assembly via the holdfast inhibitor HfiA

A genome-wide analysis of adhesion inCaulobacter crescentusidentifies new regulatory and biosynthetic components for holdfast assembly

The role ofCaulobactercell surface structures in colonization of the air-liquid interface

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