Quorum‐sensing autoinducer molecules produced by members of a multispecies biofilm promote horizontal gene transfer to Vibrio cholerae
TL;DR: It is demonstrated that comEA transcription and the horizontal acquisition of DNA by V. cholerae are induced in response to purified CAI-1 and AI-2, and also by autoinducers derived from other Vibrios co-cultured with V. Cholerae within a mixed-species biofilm, suggesting that autoinducer communication within a consortium may promote DNA exchange among VibRIos.
Abstract: Vibrio cholerae, the causative agent of cholera and a natural inhabitant of aquatic environments, regulates numerous behaviors using a quorum-sensing (QS) system conserved among many members of the marine genus Vibrio. The Vibrio QS response is mediated by two extracellular autoinducer (AI) molecules: CAI-I, which is produced only by Vibrios, and AI-2, which is produced by many bacteria. In marine biofilms on chitinous surfaces, QS-proficient V. cholerae become naturally competent to take up extracellular DNA. Because the direct role of AIs in this environmental behavior had not been determined, we sought to define the contribution of CAI-1 and AI-2 in controlling transcription of the competence gene, comEA, and in DNA uptake. In this study we demonstrated that comEA transcription and the horizontal acquisition of DNA by V. cholerae are induced in response to purified CAI-1 and AI-2, and also by autoinducers derived from other Vibrios co-cultured with V. cholerae within a mixed-species biofilm. These results suggest that autoinducer communication within a consortium may promote DNA exchange among Vibrios, perhaps contributing to the evolution of these bacterial pathogens.
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TL;DR: It is argued that recognition of these evolutionary bonds stimulates radical rethinking of key questions in evolutionary biology, which will expand the study of biological complexity beyond the usual genealogical bonds, and provide an important step to a more realistic pluralist treatment of evolutionary complexity.
Abstract: All evolutionary biologists are familiar with evolutionary units that evolve by vertical descent in a tree-like fashion in single lineages. However, many other kinds of processes contribute to evolutionary diversity. In vertical descent, the genetic material of a particular evolutionary unit is propagated by replication inside its own lineage. In what we call introgressive descent, the genetic material of a particular evolutionary unit propagates into different host structures and is replicated within these host structures. Thus, introgressive descent generates a variety of evolutionary units and leaves recognizable patterns in resemblance networks. We characterize six kinds of evolutionary units, of which five involve mosaic lineages generated by introgressive descent. To facilitate detection of these units in resemblance networks, we introduce terminology based on two notions, P3s (subgraphs of three nodes: A, B, and C) and mosaic P3s, and suggest an apparatus for systematic detection of introgressive descent. Mosaic P3s correspond to a distinct type of evolutionary bond that is orthogonal to the bonds of kinship and genealogy usually examined by evolutionary biologists. We argue that recognition of these evolutionary bonds stimulates radical rethinking of key questions in evolutionary biology (e.g., the relations among evolutionary players in very early phases of evolutionary history, the origin and emergence of novelties, and the production of new lineages). This line of research will expand the study of biological complexity beyond the usual genealogical bonds, revealing additional sources of biodiversity. It provides an important step to a more realistic pluralist treatment of evolutionary complexity.
75 citations
Cites background from "Quorum‐sensing autoinducer molecule..."
...Multispecies biofilms (59), environmental coalitions of cells and mobile genetic elements like those elements of marine cyanophages and cyanobacteria (28), and genetic exchange communities in gut microbiomes (31, 60, 61) provide examples of such multilineage clubs....
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TL;DR: A unique organization of the L. pneumophila Lqs system comprising two partially antagonistic LAI-1-responsive sensor kinases, LqsT and LqsS, which regulate distinct pools of genes implicated in pathogen-host cell interactions, competence, expression of a genomic island or production of extracellular filaments is revealed.
Abstract: Summary
Legionella pneumophila is an amoeba-resistant opportunistic pathogen that performs cell–cell communication through the signalling molecule 3-hydroxypentadecane-4-one (LAI-1, Legionella autoinducer-1). The lqs (Legionella quorum sensing) gene cluster encodes the LAI-1 autoinducer synthase LqsA, the cognate sensor kinase LqsS and the response regulator LqsR. Here we show that the Lqs system includes an ‘orphan’ homologue of LqsS termed LqsT. Compared with wild-type L. pneumophila, strains lacking lqsT or both lqsS and lqsT show increased salt resistance, greatly enhanced natural competence for DNA acquisition and impaired uptake by phagocytes. Sensitive novel single round growth assays and competition experiments using Acanthamoeba castellanii revealed that ΔlqsT and ΔlqsS-ΔlqsT, as well as ΔlqsA and other lqs mutant strains are impaired for intracellular growth and cannot compete against wild-type bacteria upon co-infection. In contrast to the ΔlqsS strain, ΔlqsT does not produce extracellular filaments. The phenotypes of the ΔlqsS-ΔlqsT strain are partially complemented by either lqsT or lqsS, but are not reversed by overexpression of lqsA, suggesting that LqsT and LqsS are the sole LAI-1-responsive sensor kinases in L. pneumophila. In agreement with the different phenotypes of the ΔlqsT and ΔlqsS strains, lqsT and lqsS are differentially expressed in the post-exponential growth phase, and transcriptome studies indicated that 90% of the genes, which are downregulated in absence of lqsT, are upregulated in absence of lqsS. Reciprocally regulated genes encode components of a 133 kb genomic ‘fitness island’ or translocated effector proteins implicated in virulence. Together, these results reveal a unique organization of the L. pneumophila Lqs system comprising two partially antagonistic LAI-1-responsive sensor kinases, LqsT and LqsS, which regulate distinct pools of genes implicated in pathogen–host cell interactions, competence, expression of a genomic island or production of extracellular filaments.
69 citations
Cites background from "Quorum‐sensing autoinducer molecule..."
...The diversity, complexity and cross-talk of these systems is almost overwhelming (Fuqua and Greenberg, 2002; Bassler and Losick, 2006; Camilli and Bassler, 2006; Winans, 2011)....
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...© 2012 Society for Applied Microbiology and Blackwell Publishing Ltd, Environmental Microbiology, 15, 646–662 Role of lqs genes in the regulation of natural competence of L. pneumophila Quorum sensing and small molecule signalling regulates natural competence of bacteria (Bassler and Losick, 2006)....
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TL;DR: High-throughput RNA sequencing shows that in V. cholerae, CytR is a global regulator of multiple behaviors affecting fitness and adaptability in the environment.
Abstract: The facultative pathogen Vibrio cholerae transitions between its human host and aquatic reservoirs where it colonizes chitinous surfaces. Growth on chitin induces expression of chitin utilization genes, genes involved in DNA uptake by natural transformation, and a type VI secretion system that allows contact-dependent killing of neighboring bacteria. We have previously shown that the transcription factor CytR, thought to primarily regulate the pyrimidine nucleoside scavenging response, is required for natural competence in V. cholerae. Through high-throughput RNA sequencing (RNA-seq), we show that CytR positively regulates the majority of competence genes, the three type VI secretion operons, and the four known or predicted chitinases. We used transcriptional reporters and phenotypic analysis to determine the individual contributions of quorum sensing, which is controlled by the transcription factors HapR and QstR; chitin utilization that is mediated by TfoX; and pyrimidine starvation that is orchestrated by CytR, toward each of these processes. We find that in V. cholerae, CytR is a global regulator of multiple behaviors affecting fitness and adaptability in the environment.
64 citations
Cites background or methods from "Quorum‐sensing autoinducer molecule..."
...3 was reached, and genomic DNA marked with a kanamycin resistance cassette [12] was then added at a final conc....
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...cholerae in response to secreted autoinducer signals at high density [4, 8, 12, 13]....
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...Expression of each reporter and of the previously published comEA-lux and pilA-lux reporters [12, 16] was measured in a V....
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TL;DR: It is demonstrated that the recently identified autoinducer, DPO, also controls AphA production in V. cholerae and that all three autoinducers are required for a full quorum sensing response.
Abstract: Bacteria use quorum sensing to monitor cell density and coordinate group behaviours. In Vibrio cholerae, the causative agent of the diarrheal disease cholera, quorum sensing is connected to virulence gene expression via the two autoinducer molecules, AI-2 and CAI-1. Both autoinducers share one signal transduction pathway to control the production of AphA, a key transcriptional activator of biofilm formation and virulence genes. In this study, we demonstrate that the recently identified autoinducer, DPO, also controls AphA production in V. cholerae. DPO, functioning through the transcription factor VqmA and the VqmR small RNA, reduces AphA levels at the post-transcriptional level and consequently inhibits virulence gene expression. VqmR-mediated repression of AphA provides an important link between the AI-2/CAI-1 and DPO-dependent quorum sensing pathways in V. cholerae. Transcriptome analyses comparing the effect of single autoinducers versus autoinducer combinations show that quorum sensing controls the expression of ∼400 genes in V. cholerae and that all three autoinducers are required for a full quorum sensing response. Together, our data provide a global view on autoinducer interplay in V. cholerae and highlight the importance of RNA-based gene control for collective functions in this major human pathogen.
64 citations
Cites background from "Quorum‐sensing autoinducer molecule..."
...In Vibrio cholerae, the causative agent of cholera disease, QS is intimately linked to several collective functions, including biofilm formation (3), type VI secretion (4,5), competence (6,7), phage resistance (8) and virulence gene expression (9)....
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TL;DR: The results bolster the pathoadaptive model in which tight regulation of T6SS-mediated bacterial killing is beneficial in a human host, whereas constitutive killing by environmental isolates may give a competitive advantage in natural settings.
Abstract: The bacterial pathogen Vibrio cholerae can occupy both the human gut and aquatic reservoirs, where it may colonize chitinous surfaces that induce the expression of factors for three phenotypes: chitin utilization, DNA uptake by natural transformation, and contact-dependent bacterial killing via a type VI secretion system (T6SS). In this study, we surveyed a diverse set of 53 isolates from different geographic locales collected over the past century from human clinical and environmental specimens for each phenotype outlined above. The set included pandemic isolates of serogroup O1, as well as several serogroup O139 and non-O1/non-O139 strains. We found that while chitin utilization was common, only 22.6% of the isolates tested were proficient at chitin-induced natural transformation, suggesting that transformation is expendable. Constitutive contact-dependent killing of Escherichia coli prey, which is indicative of a functional T6SS, was rare among clinical isolates (only 4 of 29) but common among environmental isolates (22 of 24). These results bolster the pathoadaptive model in which tight regulation of T6SS-mediated bacterial killing is beneficial in a human host, whereas constitutive killing by environmental isolates may give a competitive advantage in natural settings. Future sequence analysis of this set of diverse isolates may identify previously unknown regulators and structural components for both natural transformation and T6SS.
61 citations
Cites background or methods from "Quorum‐sensing autoinducer molecule..."
...tance gene and the tfoX gene under the control of the ptac promoter (29) was digested at the BamHI restriction enzyme site located 5= of the tfoX...
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...cholerae by upregulating the expression of transcription factor HapR in response to the accumulation of secreted autoinducer signals at high cell density (12, 21, 29, 30)....
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...0E 08 were deemed severely impaired in transformation, as described previously (29), and are designated negative in Table 1....
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References
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Book•
15 Jan 2001
TL;DR: Molecular Cloning has served as the foundation of technical expertise in labs worldwide for 30 years as mentioned in this paper and has been so popular, or so influential, that no other manual has been more widely used and influential.
Abstract: Molecular Cloning has served as the foundation of technical expertise in labs worldwide for 30 years. No other manual has been so popular, or so influential. Molecular Cloning, Fourth Edition, by the celebrated founding author Joe Sambrook and new co-author, the distinguished HHMI investigator Michael Green, preserves the highly praised detail and clarity of previous editions and includes specific chapters and protocols commissioned for the book from expert practitioners at Yale, U Mass, Rockefeller University, Texas Tech, Cold Spring Harbor Laboratory, Washington University, and other leading institutions. The theoretical and historical underpinnings of techniques are prominent features of the presentation throughout, information that does much to help trouble-shoot experimental problems. For the fourth edition of this classic work, the content has been entirely recast to include nucleic-acid based methods selected as the most widely used and valuable in molecular and cellular biology laboratories. Core chapters from the third edition have been revised to feature current strategies and approaches to the preparation and cloning of nucleic acids, gene transfer, and expression analysis. They are augmented by 12 new chapters which show how DNA, RNA, and proteins should be prepared, evaluated, and manipulated, and how data generation and analysis can be handled. The new content includes methods for studying interactions between cellular components, such as microarrays, next-generation sequencing technologies, RNA interference, and epigenetic analysis using DNA methylation techniques and chromatin immunoprecipitation. To make sense of the wealth of data produced by these techniques, a bioinformatics chapter describes the use of analytical tools for comparing sequences of genes and proteins and identifying common expression patterns among sets of genes. Building on thirty years of trust, reliability, and authority, the fourth edition of Mol
215,169 citations
TL;DR: It is evident that biofilm formation is an ancient and integral component of the prokaryotic life cycle, and is a key factor for survival in diverse environments.
Abstract: Biofilms--matrix-enclosed microbial accretions that adhere to biological or non-biological surfaces--represent a significant and incompletely understood mode of growth for bacteria. Biofilm formation appears early in the fossil record (approximately 3.25 billion years ago) and is common throughout a diverse range of organisms in both the Archaea and Bacteria lineages, including the 'living fossils' in the most deeply dividing branches of the phylogenetic tree. It is evident that biofilm formation is an ancient and integral component of the prokaryotic life cycle, and is a key factor for survival in diverse environments. Recent advances show that biofilms are structurally complex, dynamic systems with attributes of both primordial multicellular organisms and multifaceted ecosystems. Biofilm formation represents a protected mode of growth that allows cells to survive in hostile environments and also disperse to colonize new niches. The implications of these survival and propagative mechanisms in the context of both the natural environment and infectious diseases are discussed in this review.
6,170 citations
"Quorum‐sensing autoinducer molecule..." refers background in this paper
...We reasoned that a mixed-species consortium may more closely reflect conditions in environmental biofilms that are unlikely to be mono-species in composition (Hall-Stoodley et al., 2004; Wintermute & Silver, 2010)....
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TL;DR: The evolution of quorum sensing systems in bacteria could, therefore, have been one of the early steps in the development of multicellularity.
Abstract: ▪ Abstract Quorum sensing is the regulation of gene expression in response to fluctuations in cell-population density. Quorum sensing bacteria produce and release chemical signal molecules called autoinducers that increase in concentration as a function of cell density. The detection of a minimal threshold stimulatory concentration of an autoinducer leads to an alteration in gene expression. Gram-positive and Gram-negative bacteria use quorum sensing communication circuits to regulate a diverse array of physiological activities. These processes include symbiosis, virulence, competence, conjugation, antibiotic production, motility, sporulation, and biofilm formation. In general, Gram-negative bacteria use acylated homoserine lactones as autoinducers, and Gram-positive bacteria use processed oligo-peptides to communicate. Recent advances in the field indicate that cell-cell communication via autoinducers occurs both within and between bacterial species. Furthermore, there is mounting data suggesting that ba...
4,449 citations
"Quorum‐sensing autoinducer molecule..." refers background in this paper
...…molecules has been studied in many laboratory systems that relied exclusively on cell-free culture fluids or monocultures (Bassler et al., 1997; Miller & Bassler, 2001; Henke & Bassler, 2004a), single-species co-cultures (Hammer & Bassler, 2007), or cocultures of Vibrios with other bacteria…...
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TL;DR: How the marine luminescent bacterium V. fischeri uses the LuxR and LuxI proteins for intercellular communication is reviewed and a newly discovered family of LuxRand LuxI homologs in diverse bacterial species is described.
Abstract: It has long been appreciated that certain groups of bacteria exhibit cooperative behavioral patterns. For example, feeding and sporulation of both myxobacteria and actinomycetes seem optimized for large populations of cells behaving almost as a single multicellular organism. The swarming motility of microorganisms such as Vibrio parahaemolyticus and Proteus mirabilis provides another excellent example of multicellular behavior among bacteria (2). Intercellular communication likewise has been appreciated for several years in Vibrio fischeri, Myxococcus xanthus, Bacillus subtilis, Streptomyces spp., the eukaryotic slime mold Dictyostelium discoideum, and other species (44). Here we first review how the marine luminescent bacterium V. fischeri uses the LuxR and LuxI proteins for intercellular communication and then describe a newly discovered family of LuxR and LuxI homologs in diverse bacterial species.
2,693 citations
"Quorum‐sensing autoinducer molecule..." refers background in this paper
...…bacterium and the causative agent of the disease cholera, produces and then responds to extracellular small molecules called autoinducers (AIs) to collectively control gene expression and coordinate group behaviors, a process called quorum sensing (QS) (Fuqua et al., 1994; Ng & Bassler, 2009)....
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TL;DR: The emergence of toxigenic V. cholerae involves horizontal gene transfer that may depend on in vivo gene expression, and is shown here to be encoded by a filamentous bacteriophage (designated CTXΦ), which is related to coliphage M13.
Abstract: Vibrio cholerae, the causative agent of cholera, requires two coordinately regulated factors for full virulence: cholera toxin (CT), a potent enterotoxin, and toxin-coregulated pili (TCP), surface organelles required for intestinal colonization. The structural genes for CT are shown here to be encoded by a filamentous bacteriophage (designated CTXphi), which is related to coliphage M13. The CTXphi genome chromosomally integrated or replicated as a plasmid. CTXphi used TCP as its receptor and infected V. cholerae cells within the gastrointestinal tracts of mice more efficiently than under laboratory conditions. Thus, the emergence of toxigenic V. cholerae involves horizontal gene transfer that may depend on in vivo gene expression.
1,744 citations
"Quorum‐sensing autoinducer molecule..." refers background in this paper
...Transduction of the cholera toxin genes encoded within a filamentous phage (CTXF) permits exchange of virulence factors among V. cholerae (Waldor & Mekalanos, 1996)....
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