Identification and analysis of the signaling pathways, matrix-digestion enzymes, and motility components controlling Vibrio cholerae biofilm dispersal

Bacteria alternate between being free-swimming and existing as members of sessile multicellular communities called biofilms. The biofilm lifecycle occurs in three stages: cell attachment, biofilm maturation, and biofilm dispersal.Vibrio choleraebiofilms are hyperinfectious, and biofilm formation and dispersal are considered central to disease transmission. While biofilm formation is well studied, almost nothing is known about biofilm dispersal. Here, we conducted an imaging screen forV. choleraemutants that fail to disperse, revealing three classes of dispersal components: signal transduction proteins, matrix-degradation enzymes, and motility factors. Signaling proteins dominated the screen and among them, we focused on an uncharacterized two-component sensory system that we term DbfS/DbfR for dispersal of biofilm sensor/regulator. Phospho-DbfR represses biofilm dispersal. DbfS dephosphorylates and thereby inactivates DbfR, which permits dispersal. Matrix degradation requires two enzymes: LapG, which cleaves adhesins, and RbmB, which digests matrix polysaccharides. Reorientation in swimming direction, mediated by CheY3, is necessary for cells to escape from the porous biofilm matrix. We suggest that these components act sequentially: signaling launches dispersal by terminating matrix production and triggering matrix digestion, and subsequent cell motility permits escape from biofilms. This study lays the groundwork for interventions aimed at modulatingV. choleraebiofilm dispersal to ameliorate disease.

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Identification of signaling pathways, matrix-digestion enzymes, and motility components controlling Vibrio cholerae biofilm dispersal

10.1101/2020.10.09.333351 ◽

2020 ◽

Author(s):

Andrew A. Bridges ◽

Chenyi Fei ◽

Bonnie L. Bassler

Keyword(s):

Signal Transduction ◽

Vibrio Cholerae ◽

Disease Transmission ◽

Biofilm Formation ◽

Cell Attachment ◽

Sensory System ◽

Matrix Degradation ◽

Free Swimming ◽

Molecular Events ◽

Biofilm Dispersal

AbstractBacteria alternate between being free-swimming and existing as members of sessile multicellular communities called biofilms. The biofilm lifecycle occurs in three stages: cell attachment, biofilm maturation, and biofilm dispersal. Vibrio cholerae biofilms are hyper-infectious and biofilm formation and dispersal are considered central to disease transmission. While biofilm formation is well-studied, almost nothing is known about biofilm dispersal. Here, we conduct an imaging screen for V. cholerae mutants that fail to disperse, revealing three classes of dispersal components: signal transduction proteins, matrix-degradation enzymes, and motility factors. Signaling proteins dominated the screen and among them, we focused on an uncharacterized two-component sensory system that we name DbfS/DbfR for Dispersal of Biofilm Sensor/Regulator. Phospho-DbfR represses biofilm dispersal. DbfS dephosphorylates and thereby inactivates DbfR, which permits dispersal. Matrix degradation requires two enzymes: LapG, which cleaves adhesins, and RbmB, which digests matrix polysaccharide. Reorientations in swimming direction, mediated by CheY3, are necessary for cells to escape from the porous biofilm matrix. We suggest that these components act sequentially: signaling launches dispersal by terminating matrix production and triggering matrix digestion and, subsequently, cell motility permits escape from biofilms. This study lays the groundwork for interventions that modulate V. cholerae biofilm dispersal to ameliorate disease.Significance statementThe pathogen Vibrio cholerae alternates between the free-swimming state and existing in sessile multicellular communities known as biofilms. Transitioning between these lifestyles is key for disease transmission. V. cholerae biofilm formation is well studied, however, almost nothing is known about how V. cholerae cells disperse from biofilms, precluding understanding of a central pathogenicity step. Here, we conducted a high-content imaging screen for V. cholerae mutants that failed to disperse. Our screen revealed three classes of components required for dispersal: signal transduction, matrix degradation, and motility factors. We characterized these components to reveal the sequence of molecular events that choreograph V. cholerae biofilm dispersal. Our report provides a framework for developing strategies to modulate biofilm dispersal to prevent or treat disease.

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Toxin-Antitoxin Systems in Escherichia coli Influence Biofilm Formation through YjgK (TabA) and Fimbriae

Journal of Bacteriology ◽

10.1128/jb.01465-08 ◽

2008 ◽

Vol 191 (4) ◽

pp. 1258-1267 ◽

Cited By ~ 120

Author(s):

Younghoon Kim ◽

Xiaoxue Wang ◽

Qun Ma ◽

Xue-Song Zhang ◽

Thomas K. Wood

Keyword(s):

Biofilm Formation ◽

Cell Attachment ◽

Single Gene ◽

Transcriptome Profiling ◽

Uncharacterized Protein ◽

Qrt Pcr ◽

Biofilm Dispersal ◽

Biofilm Development ◽

Whole Transcriptome

ABSTRACT The roles of toxin-antitoxin (TA) systems in bacteria have been debated. Here, the role of five TA systems in regard to biofilm development was investigated (listed as toxin/antitoxin: MazF/MazE, RelE/RelB, ChpB, YoeB/YefM, and YafQ/DinJ). Although these multiple TA systems were reported previously to not impact bacterial fitness, we found that deletion of the five TA systems decreased biofilm formation initially (8 h) on three different surfaces and then increased biofilm formation (24 h) by decreasing biofilm dispersal. Whole-transcriptome profiling revealed that the deletion of the five TA systems induced expression of a single gene, yjgK, which encodes an uncharacterized protein; quantitative real-time PCR (qRT-PCR) confirmed consistent induction of this gene (at 8, 15, and 24 h). Corroborating the complex phenotype seen upon deleting the TA systems, overexpression of YjgK decreased biofilm formation at 8 h and increased biofilm formation at 24 h; deletion of yjgK also affected biofilm formation in the expected manner by increasing biofilm formation after 8 h and decreasing biofilm formation after 24 h. In addition, YjgK significantly reduced biofilm dispersal. Whole-transcriptome profiling revealed YjgK represses fimbria genes at 8 h (corroborated by qRT-PCR and a yeast agglutination assay), which agrees with the decrease in biofilm formation upon deleting the five TA systems at 8 h, as well as that seen upon overexpressing YjgK. Sand column assays confirmed that deleting the five TA systems reduced cell attachment. Furthermore, deletion of each of the five toxins increased biofilm formation at 8 h, and overexpression of the five toxins repressed biofilm formation at 8 h, a result that is opposite that of deleting all five TA systems; this suggests that complex regulation occurs involving the antitoxins. Also, the ability of the global regulator Hha to reduce biofilm formation was dependent on the presence of these TA systems. Hence, we suggest that one role of TA systems is to influence biofilm formation.

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A Conserved Regulatory Circuit Controls Large Adhesins in Vibrio cholerae

mBio ◽

10.1128/mbio.02822-19 ◽

2019 ◽

Vol 10 (6) ◽

Cited By ~ 6

Author(s):

Giordan Kitts ◽

Krista M. Giglio ◽

David Zamorano-Sánchez ◽

Jin Hwan Park ◽

Loni Townsley ◽

...

Keyword(s):

Cell Surface ◽

Vibrio Cholerae ◽

Biofilm Formation ◽

Diarrheal Disease ◽

Cell Attachment ◽

Surface Display ◽

Second Messenger ◽

Bacterial Biofilm ◽

Content Type ◽

Regulatory Circuit

ABSTRACT The dinucleotide second messenger c-di-GMP has emerged as a central regulator of reversible cell attachment during bacterial biofilm formation. A prominent cell adhesion mechanism first identified in pseudomonads combines two c-di-GMP-mediated processes: transcription of a large adhesin and its cell surface display via posttranslational proteolytic control. Here, we characterize an orthologous c-di-GMP effector system and show that it is operational in Vibrio cholerae, where it regulates two distinct classes of adhesins. Through structural analyses, we reveal a conserved autoinhibition mechanism of the c-di-GMP receptor that controls adhesin proteolysis and present a structure of a c-di-GMP-bound receptor module. We further establish functionality of the periplasmic protease controlled by the receptor against the two adhesins. Finally, transcription and functional assays identify physiological roles of both c-di-GMP-regulated adhesins in surface attachment and biofilm formation. Together, our studies highlight the conservation of a highly efficient signaling effector circuit for the control of cell surface adhesin expression and its versatility by revealing strain-specific variations. IMPORTANCE Vibrio cholerae, the causative agent of the diarrheal disease cholera, benefits from a sessile biofilm lifestyle that enhances survival outside the host but also contributes to host colonization and infectivity. The bacterial second messenger c-di-GMP has been identified as a central regulator of biofilm formation, including in V. cholerae; however, our understanding of the pathways that contribute to this process is incomplete. Here, we define a conserved signaling system that controls the stability of large adhesion proteins at the cell surface of V. cholerae, which are important for cell attachment and biofilm formation. Insight into the regulatory circuit underlying biofilm formation may inform targeted strategies to interfere with a process that renders this bacterium remarkably adaptable to changing environments.

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Inverse regulation of Vibrio cholerae biofilm dispersal by polyamine signals

10.1101/2020.12.06.413526 ◽

2020 ◽

Author(s):

Andrew A. Bridges ◽

Bonnie L. Bassler

Keyword(s):

Vibrio Cholerae ◽

Biofilm Formation ◽

Self And Other ◽

Human Host ◽

Signal Relay ◽

Biofilm Dispersal ◽

Methylene Group

AbstractThe global pathogen Vibrio cholerae undergoes cycles of biofilm formation and dispersal in the environment and the human host. Little is understood about biofilm dispersal. Here, we show that MbaA, a periplasmic polyamine sensor, and PotD1, a polyamine importer, regulate V. cholerae biofilm dispersal. Spermidine, a commonly produced polyamine, drives V. cholerae dispersal, whereas norspermidine, an uncommon polyamine produced by vibrios, inhibits dispersal. Spermidine and norspermidine differ by one methylene group. Both polyamines function to control dispersal via periplasmic detection by MbaA and subsequent signal relay. Biofilm dispersal fails in the absence of PotD1 because reuptake of endogenously produced norspermidine does not occur, so it accumulates in the periplasm where it stimulates MbaA. These results suggest that V. cholerae uses MbaA to monitor environmental polyamines, blends of which potentially provide information about numbers of ‘self’ and ‘other’. This information is used to dictate whether or not to disperse from biofilms.

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Inverse regulation of Vibrio cholerae biofilm dispersal by polyamine signals

eLife ◽

10.7554/elife.65487 ◽

2021 ◽

Vol 10 ◽

Author(s):

Andrew A Bridges ◽

Bonnie L Bassler

Keyword(s):

Vibrio Cholerae ◽

Biofilm Formation ◽

Self And Other ◽

Human Host ◽

Signal Relay ◽

Biofilm Dispersal ◽

Methylene Group

The global pathogen Vibrio cholerae undergoes cycles of biofilm formation and dispersal in the environment and the human host. Little is understood about biofilm dispersal. Here, we show that MbaA, a periplasmic polyamine sensor, and PotD1, a polyamine importer, regulate V. cholerae biofilm dispersal. Spermidine, a commonly produced polyamine, drives V. cholerae dispersal, whereas norspermidine, an uncommon polyamine produced by vibrios, inhibits dispersal. Spermidine and norspermidine differ by one methylene group. Both polyamines control dispersal via MbaA detection in the periplasm and subsequent signal relay. Our results suggest that dispersal fails in the absence of PotD1 because endogenously produced norspermidine is not reimported, periplasmic norspermidine accumulates, and it stimulates MbaA signaling. These results suggest that V. cholerae uses MbaA to monitor environmental polyamines, blends of which potentially provide information about numbers of ‘self’ and ‘other’. This information is used to dictate whether or not to disperse from biofilms.

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Optimized quinoline amino alcohols as disruptors and dispersal agents of Vibrio cholerae biofilms

Organic & Biomolecular Chemistry ◽

10.1039/c5ob01134e ◽

2015 ◽

Vol 13 (31) ◽

pp. 8495-8499 ◽

Cited By ~ 6

Author(s):

Brian León ◽

F. P. Jake Haeckl ◽

Roger G. Linington

Keyword(s):

Vibrio Cholerae ◽

Amino Alcohol ◽

Biofilm Formation ◽

Bacterial Pathogens ◽

Amino Alcohols ◽

Human Pathogen ◽

Biofilm Dispersal ◽

Dispersal Agents

The biofilm state is an integral part of the lifecycle of many bacterial pathogens, but no treatments currently exist that directly impact biofilm formation or persistence. Here we report the development of a quinoline amino alcohol scaffold with both biofilm inhibitory and biofilm dispersal activities against the human pathogen Vibrio cholerae.

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The Histone-Like Nucleoid Structuring Protein (H-NS) Is a Repressor of Vibrio cholerae Exopolysaccharide Biosynthesis (vps) Genes

Applied and Environmental Microbiology ◽

10.1128/aem.07629-11 ◽

2012 ◽

Vol 78 (7) ◽

pp. 2482-2488 ◽

Cited By ~ 30

Author(s):

Hongxia Wang ◽

Julio C. Ayala ◽

Anisia J. Silva ◽

Jorge A. Benitez

Keyword(s):

Vibrio Cholerae ◽

Disease Transmission ◽

Aquatic Environment ◽

Biofilm Formation ◽

Content Type ◽

Exopolysaccharide Biosynthesis

ABSTRACTThe capacity ofVibrio choleraeto form biofilms has been shown to enhance its survival in the aquatic environment and play important roles in pathogenesis and disease transmission. In this study, we demonstrated that the histone-like nucleoid structuring protein is a repressor of exopolysaccharide (vps) biosynthesis genes and biofilm formation.

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