scholarly journals Regulation of Cyclic Lipopeptide Biosynthesis in Pseudomonas fluorescens by the ClpP Protease

2008 ◽  
Vol 191 (6) ◽  
pp. 1910-1923 ◽  
Author(s):  
I. de Bruijn ◽  
J. M. Raaijmakers
Keyword(s):  
Pseudomonas Fluorescens ◽  
Biofilm Formation ◽  
Site Directed Mutagenesis ◽  
Transcriptional Level ◽  
Two Component System ◽  
Swarming Motility ◽  
Cyclic Lipopeptides ◽  
Peptide Synthetases ◽  
Cyclic Lipopeptide ◽  
Casamino Acids

ABSTRACT Cyclic lipopeptides produced by Pseudomonas species exhibit potent surfactant and broad-spectrum antibiotic properties. Their biosynthesis is governed by large multimodular nonribosomal peptide synthetases, but little is known about the genetic regulatory network. This study provides, for the first time, evidence that the serine protease ClpP regulates the biosynthesis of massetolides, cyclic lipopeptides involved in swarming motility, biofilm formation, and antimicrobial activity of Pseudomonas fluorescens SS101. The results show that ClpP affects the expression of luxR(mA), the transcriptional regulator of the massetolide biosynthesis genes massABC, thereby regulating biofilm formation and swarming motility of P. fluorescens SS101. Transcription of luxR(mA) was significantly repressed in the clpP mutant, and introduction of luxR(mA) restored, in part, massetolide biosynthesis and swarming motility of the clpP mutant. Site-directed mutagenesis and expression analyses indicated that the chaperone subunit ClpX and the Lon protease are not involved in regulation of massetolide biosynthesis and are transcribed independently of clpP. Addition of Casamino Acids enhanced the transcription of luxR(mA) and massABC in the clpP mutant, leading to a partial rescue of massetolide production and swarming motility. The results further suggested that, at the transcriptional level, ClpP-mediated regulation of massetolide biosynthesis operates independently of regulation by the GacA/GacS two-component system. The role of amino acid metabolism and the putative mechanisms underlying ClpP-mediated regulation of cyclic lipopeptide biosynthesis, swarming motility, and growth in P. fluorescens are discussed.

scholarly journals Association of Hemolytic Activity of Pseudomonas entomophila, a Versatile Soil Bacterium, with Cyclic Lipopeptide Production

2009 ◽  
Vol 76 (3) ◽  
pp. 910-921 ◽  
Author(s):  
Isabelle Vallet-Gely ◽  
Alexey Novikov ◽  
Luis Augusto ◽  
Peter Liehl ◽  
Gérard Bolbach ◽  
...  
Keyword(s):  
Hemolytic Activity ◽  
Physiological Role ◽  
Soil Bacterium ◽  
Two Component System ◽  
Swarming Motility ◽  
Component System ◽  
Peptide Synthetases ◽  
Cyclic Lipopeptide ◽  
Two Component

ABSTRACT Pseudomonas entomophila is an entomopathogenic bacterium that is able to infect and kill Drosophila melanogaster upon ingestion. Its genome sequence suggests that it is a versatile soil bacterium closely related to Pseudomonas putida. The GacS/GacA two-component system plays a key role in P. entomophila pathogenicity, controlling many putative virulence factors and AprA, a secreted protease important to escape the fly immune response. P. entomophila secretes a strong diffusible hemolytic activity. Here, we showed that this activity is linked to the production of a new cyclic lipopeptide containing 14 amino acids and a 3-C10OH fatty acid that we called entolysin. Three nonribosomal peptide synthetases (EtlA, EtlB, EtlC) were identified as responsible for entolysin biosynthesis. Two additional components (EtlR, MacAB) are necessary for its production and secretion. The P. entomophila GacS/GacA two-component system regulates entolysin production, and we demonstrated that its functioning requires two small RNAs and two RsmA-like proteins. Finally, entolysin is required for swarming motility, as described for other lipopeptides, but it does not participate in the virulence of P. entomophila for Drosophila. While investigating the physiological role of entolysin, we also uncovered new phenotypes associated with P. entomophila, including strong biocontrol abilities.

scholarly journals Quorum sensing inhibitory effects of vanillin on the biofilm formation of Pseudomonas fluorescens P07 by transcriptome analysis

2021 ◽  
Vol 5 (7) ◽  
pp. 275-292
Author(s):  
Ting Ding ◽  
◽  
Yong Li
Keyword(s):  
Quorum Sensing ◽  
Pseudomonas Fluorescens ◽  
Transcriptome Analysis ◽  
Biofilm Formation ◽  
Homoserine Lactone ◽  
Bacterial Biofilm ◽  
Inhibitory Effects ◽  
Swarming Motility ◽  
Physiological Processes ◽  
Spoilage Bacterium

Pseudomonas fluorescens is an important psychrotrophic food-spoilage bacterium. Quorum sensing (QS) enables bacteria to control various physiological processes. Hence, targeting bacterial QS would be a novel method to improve food quality. In this study, P. fluorescens P07 was treated with vanillin, which showed strong QS inhibitory activity, and its resultant effects on swarming motility, biofilm formation, and extracellular polymeric substance (EPS) secretion were measured. The mechanisms underlying the inhibitory effects were then explored by transcriptomic analysis. The results showed that vanillin had inhibitory effects on swarming motility, biofilm formation, N-acyl-L-homoserine Lactone (AHLs) and EPS secretion of P. fluorescens P07. The result of transcriptionomic tests indicated that the decrease in bacterial biofilm formation was probably due to the influence of vanillin on mobility, adhesion, chemotaxis, EPS secretion, and QS system of the bacteria. Keywords: Pseudomonas fluorescens, quorum sensing, biofilm formation, transcriptome analysis, swarming motility

scholarly journals The QseEF Two-Component System-GlmY Small RNA Regulatory Pathway Controls Swarming in Uropathogenic Proteus mirabilis

2022 ◽  
Vol 23 (1) ◽  
pp. 487
Author(s):  
Wen-Yuan Lin ◽  
Yuan-Ju Lee ◽  
Ping-Hung Yu ◽  
Yi-Lin Tsai ◽  
Pin-Yi She ◽  
...  
Keyword(s):  
Small Rna ◽  
Proteus Mirabilis ◽  
Promoter Activity ◽  
Mrna Level ◽  
Site Directed Mutagenesis ◽  
Dependent Manner ◽  
Two Component System ◽  
Swarming Motility ◽  
Component System ◽  
Two Component

Bacterial sensing of environmental signals through the two-component system (TCS) plays a key role in modulating virulence. In the search for the host hormone-sensing TCS, we identified a conserved qseEGF locus following glmY, a small RNA (sRNA) gene in uropathogenic Proteus mirabilis. Genes of glmY-qseE-qseG-qseF constitute an operon, and QseF binding sites were found in the glmY promoter region. Deletion of glmY or qseF resulted in reduced swarming motility and swarming-related phenotypes relative to the wild-type and the respective complemented strains. The qseF mutant had decreased glmYqseEGF promoter activity. Both glmY and qseF mutants exhibited decreased flhDC promoter activity and mRNA level, while increased rcsB mRNA level was observed in both mutants. Prediction by TargetRNA2 revealed cheA as the target of GlmY. Then, construction of the translational fusions containing various lengths of cheA 5′UTR for reporter assay and site-directed mutagenesis were performed to investigate the cheA-GlmY interaction in cheA activation. Notably, loss of glmY reduced the cheA mRNA level, and urea could inhibit swarming in a QseF-dependent manner. Altogether, this is the first report elucidating the underlying mechanisms for modulation of swarming motility by a QseEF-regulated sRNA GlmY, involving expression of cheA, rcsB and flhDC in uropathogenic P. mirabilis.

scholarly journals Massetolide A Biosynthesis in Pseudomonas fluorescens

2007 ◽  
Vol 190 (8) ◽  
pp. 2777-2789 ◽  
Author(s):  
I. de Bruijn ◽  
M. J. D. de Kock ◽  
P. de Waard ◽  
T. A. van Beek ◽  
J. M. Raaijmakers
Keyword(s):  
In Silico Analysis ◽  
Gene Clusters ◽  
Site Directed Mutagenesis ◽  
Pcr Analysis ◽  
Exponential Growth Phase ◽  
Swarming Motility ◽  
Acylhomoserine Lactone ◽  
Cyclic Lipopeptide ◽  
Peptide Moiety ◽  
Peptide Synthetase

ABSTRACT Massetolide A is a cyclic lipopeptide (CLP) antibiotic produced by various Pseudomonas strains from diverse environments. Cloning, sequencing, site-directed mutagenesis, and complementation showed that massetolide A biosynthesis in P. fluorescens SS101 is governed by three nonribosomal peptide synthetase (NRPS) genes, designated massA, massB, and massC, spanning approximately 30 kb. Prediction of the nature and configuration of the amino acids by in silico analysis of adenylation and condensation domains of the NRPSs was consistent with the chemically determined structure of the peptide moiety of massetolide A. Structural analysis of massetolide A derivatives produced by SS101 indicated that most of the variations in the peptide moiety occur at amino acid positions 4 and 9. Regions flanking the mass genes contained several genes found in other Pseudomonas CLP biosynthesis clusters, which encode LuxR-type transcriptional regulators, ABC transporters, and an RND-like outer membrane protein. In contrast to most Pseudomonas CLP gene clusters known to date, the mass genes are not physically linked but are organized in two separate clusters, with massA disconnected from massB and massC. Quantitative real-time PCR analysis indicated that transcription of massC is strongly reduced when massB is mutated, suggesting that these two genes function in an operon, whereas transcription of massA is independent of massBC and vice versa. Massetolide A is produced in the early exponential growth phase, and biosynthesis appears not to be regulated by N-acylhomoserine lactone-based quorum sensing. Massetolide A production is essential in swarming motility of P. fluorescens SS101 and plays an important role in biofilm formation.

scholarly journals Biochemical, Genetic, and Zoosporicidal Properties of Cyclic Lipopeptide Surfactants Produced by Pseudomonas fluorescens

2003 ◽  
Vol 69 (12) ◽  
pp. 7161-7172 ◽  
Author(s):  
Jorge T. de Souza ◽  
Marjan de Boer ◽  
Pieter de Waard ◽  
Teris A. van Beek ◽  
Jos M. Raaijmakers
Keyword(s):  
Surface Tension ◽  
Pseudomonas Fluorescens ◽  
High Performance ◽  
Cell Suspensions ◽  
Main Constituent ◽  
Peptide Synthetases ◽  
Cyclic Lipopeptide ◽  
Biochemical Genetic ◽  
Culture Supernatants

ABSTRACT Zoospores play an important role in the infection of plant and animal hosts by oomycetes and other zoosporic fungi. In this study, six fluorescent Pseudomonas isolates with zoosporicidal activities were obtained from the wheat rhizosphere. Zoospores of multiple oomycetes, including Pythium species, Albugo candida, and Phytophthora infestans, were rendered immotile within 30 s of exposure to cell suspensions or cell culture supernatants of the six isolates, and subsequent lysis occurred within 60 s. The representative strain SS101, identified as Pseudomonas fluorescens biovar II, reduced the surface tension of water from 73 to 30 mN m−1. The application of cell suspensions of strain SS101 to soil or hyacinth bulbs provided significant protection against root rot caused by Pythium intermedium. Five Tn5 mutants of strain SS101lacked the abilities to reduce the surface tension of water and to cause lysis of zoospores. Genetic characterization of two surfactant-deficient mutants showed that the transposons had integrated into condensation domains of peptide synthetases. A partially purified extract from strain SS101 reduced the surface tension of water to 30 mN m−1 and reached the critical micelle concentration at 25 μg ml−1. Reverse-phase high-performance liquid chromatography yielded eight different fractions, five of which had surface activity and caused lysis of zoospores. Mass spectrometry and nuclear magnetic resonance analyses allowed the identification of the main constituent as a cyclic lipopeptide (1,139 Da) containing nine amino acids and a 10-carbon hydroxy fatty acid. The other four zoosporicidal fractions were closely related to the main constituent, with molecular massesranging from 1,111 to 1,169 Da.

scholarly journals Diversity and Functional Analysis of LuxR-Type Transcriptional Regulators of Cyclic Lipopeptide Biosynthesis in Pseudomonas fluorescens

2009 ◽  
Vol 75 (14) ◽  
pp. 4753-4761 ◽  
Author(s):  
I. de Bruijn ◽  
J. M. Raaijmakers
Keyword(s):  
Pseudomonas Fluorescens ◽  
Response Regulator ◽  
Phylogenetic Analyses ◽  
Regulatory Gene ◽  
Transcriptional Regulators ◽  
Site Directed Mutagenesis ◽  
Acylhomoserine Lactone ◽  
Cyclic Lipopeptide ◽  
Surface Motility

ABSTRACT Cyclic lipopeptides (CLPs) are produced by many Pseudomonas species and have several biological functions, including a role in surface motility, biofilm formation, virulence, and antimicrobial activity. This study focused on the diversity and role of LuxR-type transcriptional regulators in CLP biosynthesis in Pseudomonas species and, specifically, viscosin production by Pseudomonas fluorescens strain SBW25. Phylogenetic analyses showed that CLP biosynthesis genes in Pseudomonas strains are flanked by LuxR-type regulators that contain a DNA-binding helix-turn-helix domain but lack N-acylhomoserine lactone-binding or response regulator domains. For SBW25, site-directed mutagenesis of the genes coding for either of the two identified LuxR-type regulators, designated ViscAR and ViscBCR, strongly reduced transcript levels of the viscABC biosynthesis genes and resulted in a loss of viscosin production. Expression analyses further showed that a mutation in either viscAR or viscBCR did not substantially (change of <2.5-fold) affect transcription of the other regulator. Transformation of the ΔviscAR mutant of SBW25 with a LuxR-type regulatory gene from P. fluorescens strain SS101 that produces massetolide, a CLP structurally related to viscosin, restored transcription of the viscABC genes and viscosin production. The results further showed that a functional viscAR gene was required for heterologous expression of the massetolide biosynthesis genes of strain SS101 in strain SBW25, leading to the production of both viscosin and massetolide. Collectively, these results indicate that the regulators flanking the CLP biosynthesis genes in Pseudomonas species represent a unique LuxR subfamily of proteins and that viscosin biosynthesis in P. fluorescens SBW25 is controlled by two LuxR-type transcriptional regulators.

scholarly journals Amplifying and Fine-Tuning Rsm sRNAs Expression and Stability to Optimize the Survival of Pseudomonas brassicacerum in Nutrient-Poor Environments

2021 ◽  
Vol 9 (2) ◽  
pp. 250
Author(s):  
David Lalaouna ◽  
Sylvain Fochesato ◽  
Mourad Harir ◽  
Philippe Ortet ◽  
Philippe Schmitt-Kopplin ◽  
...  
Keyword(s):  
Biofilm Formation ◽  
Plant Root ◽  
Stringent Response ◽  
Fine Tuning ◽  
Transcriptional Level ◽  
Two Component System ◽  
Encoding Gene ◽  
The Stability ◽  
Encoding Genes ◽  
Genetic Amplification

In the beneficial plant root-associated Pseudomonas brassicacearum strain NFM421, the GacS/GacA two-component system positively controls biofilm formation and the production of secondary metabolites through the synthesis of rsmX, rsmY and rsmZ. Here, we evidenced the genetic amplification of Rsm sRNAs by the discovery of a novel 110-nt long sRNA encoding gene, rsmX-2, generated by the duplication of rsmX-1 (formerly rsmX). Like the others rsm genes, its overexpression overrides the gacA mutation. We explored the expression and the stability of rsmX-1, rsmX-2, rsmY and rsmZ encoding genes under rich or nutrient-poor conditions, and showed that their amount is fine-tuned at the transcriptional and more interestingly at the post-transcriptional level. Unlike rsmY and rsmZ, we noticed that the expression of rsmX-1 and rsmX-2 genes was exclusively GacA-dependent. The highest expression level and longest half-life for each sRNA were correlated with the highest ppGpp and cyclic-di-GMP levels and were recorded under nutrient-poor conditions. Together, these data support the view that the Rsm system in P. brassicacearum is likely linked to the stringent response, and seems to be required for bacterial adaptation to nutritional stress.

scholarly journals Cyclic Lipopeptides with Herbicidal and Insecticidal Activities Produced by Bacillus clausii DTM1

2015 ◽  
Vol 10 (12) ◽  
pp. 1934578X1501001 ◽  
Author(s):  
Da-Le Guo ◽  
Bo Wan ◽  
Shi-Ji Xiao ◽  
Sarah Allen ◽  
Yu-Cheng Gu ◽  
...  
Keyword(s):  
Methyl Ester ◽  
Natural Product ◽  
Fermentation Broth ◽  
Cyclic Lipopeptides ◽  
Bacillus Clausii ◽  
Cyclic Lipopeptide ◽  
Lipopeptide Biosurfactants ◽  
First Time ◽  
Insecticidal Activities

Seven cyclic lipopeptide biosurfactants (1–7) were isolated for the first time from the fermentation broth of endophytic Bacillus clausii DTM1 and were identified as anteisoC13[Val7] surfactin-(L-Glu)-O-methyl-ester (1), anteisoC12[Val7] surfactin (2), anteisoC15[Val7] surfactin (3), isoC14[Leu7] surfactin (4), anteisoC12[Leu7] surfactin (5), nC13[Leu7] surfactin (6), and anteisoC14[Leu7] surfactin-(L-Glu)-O-methyl-ester (7); 1 has not been isolated before as a natural product from any source. Plate-based herbicide and insecticide bioassays showed that all compounds exhibited interesting insecticidal and herbicidal activities.

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