plasminogen-activator-inhibitor-2 and 1-phenazinecarboxylic-acid

A number of gram-negative bacteria have a quorum-sensing system and produce the N-acylhomoserine lactone (AHL) as a signal molecule. Pseudomonas chlororaphis subsp. aurantiaca StFRB508 produces one of the phenazine derivatives, phenazine-1-carboxylic acid (PCA). Whole-genome sequencing of StFRB508 revealed the presence of two sets of AHL-synthase and AHL-receptor gene, phzIR and aurIR. The mutation of phzI drastically decreased PCA production, but the mutation of aurI did not affect PCA production. The phzI and aurI double mutant did not show any PCA production. StFRB508 produces three major AHLs, N-butyryl-L-homoserine lactone (C4-HSL), N-hexanoyl-L-homoserine lactone (C6-HSL), and N-(3-hydroxyhexanoyl)-L-homoserine lactone (3-hydroxy-C6-HSL). As the results of TLC analysis, PhzI mainly catalyzes the biosynthesis of 3-hydroxy-C6-HSL, and AurI catalyzes the biosynthesis of C4-HSL and C6-HSL. PCA production in the phzI and aurI double mutant was restored by exogenous AHLs and the most active AHL was 3-hydroxy-C6-HSL. StFRB508 showed high inhibitory activity of the development of mycelia of plant pathogenic fungi, Fusarium oxysporum f. sp. conglutinans. However, the phzI and aurI double mutant could not inhibit the development of mycelia. These results demonstrated that the multiple quorum-sensing system play an important role in PCA production and antifungal activity in StFRB508.

Topics: 4-Butyrolactone; Anti-Bacterial Agents; Antifungal Agents; Bacterial Proteins; Computer Simulation; Fusarium; Genome, Bacterial; Phenazines; Pseudomonas; Quorum Sensing

Pseudomonas sp. M18 is a rhizosphere isolate capable of producing two kinds of antifungal agents: phenazine-1-carboxylic acid (PCA) and pyoluteorin. Recently, the two well-studied quorum sensing (QS) systems of Pseudomonas aeruginosa, LasR/LasI and RhlR/RhlI, have also been identified in this strain. However, in this study, through the use of lacZ translational fusion expression analysis and acyl-homoserine lactone thin-layer chromatography (TLC) bioassays, we clearly display a more complex and distinctive hierarchy of the las and rhl QS systems in strain M18. In this QS cascade, expression of rhlI was negatively controlled by the LasR/LasI QS system. In contrast with lasI, which negatively regulated the rhlR induction, lasR exerted a positive influence on rhlR expression during the log-phase. This interrelationship indicated that the response regulators (LasR and RhlR) of the QS system are expressed independently of their cognate synthases (LasI and RhlI). Furthermore, the las system also modulated the timing and magnitude of the rhlI and rhlR maximal expression. In addition, our data imply that the lasR gene exerts its negative control on PCA production through modulation of rhlI expression. Thus, interactions between the two QS systems are strain specific.

Topics: 4-Butyrolactone; Bacterial Proteins; Chromatography, Thin Layer; Gene Expression Regulation, Bacterial; Ligases; Phenazines; Pseudomonas; Pseudomonas aeruginosa; Quorum Sensing; Transcription Factors; Virulence Factors

At least 5 kinds of N-acyl-homoserine lactones(AHLs) were identified in Pseudomonas sp. M18. They were: N-butyryl-L-homoserine lactone(C4-HSL, BHL), N-hexanoyl-L-homoserine lactone(C6-HSL, HHL), N-(3-oxohexanoyl)-L-homoserine lactone(3-Oxo-C6-HSL, OHHL), N-(3-oxooctanoyl)-L-homoserine lactone (3-Oxo-C8-HSL, OOHL) and N-(3-oxodecanoyl)-L-homoserine lactone(3-Oxo-C10-HSL, ODHL). Compared with the wide-type strain M18, the variety of the AHLs in the gacA mutant strain M18G reduced to 4 species with the decreased quantity. But the phenazine-1-carboxylic acid (PCA) production was increased by about 2-fold. The product of rhll plays an important role in synthesizing BHL and HHL. The rhll'-'lacZ translational fusion expression plasmid pMEIZ was constructed in vector pME6015 and then was introduced into the wild-type strain M18 and the gacA mutant strain M18G. The galactosidase activity in chromosomal gacA disruption mutant was only 60% of that in the wild-type strain M18. This result suggested that GacA could regulate the rhll expression positively. There was no influence on PCA production by adding exogenous BHL and HHL and both together to the culture of strains M18 and M18G. This result suggested that there was no relation between GacA's negative regulation on PCA production and positive regulation on BHL and HHL synthesis.

Topics: 4-Butyrolactone; Acyl-Butyrolactones; Bacterial Proteins; Chromatography, Thin Layer; Down-Regulation; Lac Operon; Ligases; Phenazines; Protein Biosynthesis; Pseudomonas; Up-Regulation