Utilization of Acyl-Homoserine Lactone Quorum Signals for Growth by a Soil Pseudomonad and Pseudomonas aeruginosa PAO1

Abstract

Acyl-homoserine lactones (AHLs) are employed by several Proteobacteria as quorum-sensing signals. Past studies have established that these compounds are subject to biochemical decay and can be used as growth nutrients. Here we describe the isolation of a soil bacterium, Pseudomonas strain PAI-A, that degrades 3-oxododecanoyl-homoserine lactone (3OC12HSL) and other long-acyl, but not short-acyl, AHLs as sole energy sources for growth. The small-subunit rRNA gene from strain PAI-A was 98.4% identical to that of Pseudomonas aeruginosa, but the soil isolate did not produce obvious pigments or AHLs or grow under denitrifying conditions or at 42°C. The quorum-sensing bacterium P. aeruginosa, which produces both 3OC12HSL and C4HSL, was examined for the ability to utilize AHLs for growth. It did so with a specificity similar to that of strain PAI-A, i.e., degrading long-acyl but not short-acyl AHLs. In contrast to the growth observed with strain PAI-A, P. aeruginosa strain PAO1 growth on AHLs commenced only after extremely long lag phases. Liquidchromatography-atmospheric pressure chemical ionization-mass spectrometry analyses indicate that strain PAO1 degrades long-acyl AHLs via an AHL acylase and a homoserine-generating HSL lactonase. A P. aeruginosa gene, pvdQ (PA2385), has previously been identified as being a homologue of the AHL acylase described as occurring in a Ralstonia species. Escherichia coli expressing pvdQ catalyzed the rapid inactivation of long-acyl AHLs and the release of HSL. P. aeruginosa engineered to constitutively express pvdQ did not accumulate its 3OC12HSL quorum signal when grown in rich media. However, pvdQ knockout mutants of P. aeruginosa were still able to grow by utilizing 3OC12HSL. To our knowledge, this is the first report of the degradation of AHLs by pseudomonads or other ␥-Proteobacteria, of AHL acylase activity in a quorum-sensing bacterium, of HSL lactonase activity in any bacterium, and of AHL degradation with specificity only towards AHLs with long side chains.Many bacterial species control and modulate their physiology in response to increases in their population densities in a process known as quorum sensing (reviewed in references 11 and 25). Several dozen species of Proteobacteria use acyl-homoserine lactones (AHLs) as dedicated signal molecules in this process. A diversity of AHL structures and the enzymes and proteins involved in their synthesis and recognition have been elucidated (12,27,31,35). One of the best studied quorum-sensing species is the opportunistic pathogen Pseudomonas aeruginosa, which makes and responds to two distinct AHLs: 3-oxododecanoyl-homoserine lactone (3OC12HSL, also known as PAI, the signal component of the Pseudomonas las quorum-sensing system), and butanoyl-HSL (C4HSL, also known as PAI-2, the autoinducer of the rhl quorum-sensing system). The two quorum circuits are known to control several physiologies and virulence factors associated with the infection of immunocompromised individuals and those with cystic fibrosis (40). Recently, the influence...

Exploiting Quorum Sensing To Confuse Bacterial Pathogens

“…Interestingly, homologues of AiiD were also found in various other Ralstonia and Pseudomonas spp. (46), and these bacteria were later confirmed to also possess AHL acylase activity (42,44,45,47,51,52). Biochemical analysis of the fate of 3OC10HSL incubated with purified AiiD protein confirmed that AiiD functions as an AHL acylase, releasing HSL and 3-oxodecanoic acid as the major degradation products (46).…”

“…The first gammaproteobacteria reported to have AHL aclyase activity were the soil Pseudomonas isolate PAI-A and P. aerguinosa PAO1, both of which were shown to be able to utilize AHLs with acyl chains of at least 8 carbons for growth, although with differing kinetics (42). Sequence analysis revealed that the P. aeruginosa PAO1 genome encodes four homologues predicted to belong to the Ntn hydrolase superfamily, and thus far three of these (PvdQ, QuiP, and HacB) have been functionally characterized (43).…”

“…Sequence analysis revealed that the P. aeruginosa PAO1 genome encodes four homologues predicted to belong to the Ntn hydrolase superfamily, and thus far three of these (PvdQ, QuiP, and HacB) have been functionally characterized (43). PvdQ (PA2385) is an AHL aclyase with specificity for long-chain AHLs (42,44), and crystallization studies revealed that AHL hydrolysis proceeds by a mechanism typical of the Ntn hydrolase superfamily (41). The observation that a pvdQ mutant of PAO1 retained its ability to grow on longchain AHLs suggested that other AHL-degrading enzymes were also employed by P. aeruginosa (42), and additional studies confirmed that two other Ntn hydrolase homologues, QuiP (PA1032) and HacB (PA0305), also function as AHL acylases with substrate specificity similar to that of PvdQ (45,52).…”

“…PvdQ (PA2385) is an AHL aclyase with specificity for long-chain AHLs (42,44), and crystallization studies revealed that AHL hydrolysis proceeds by a mechanism typical of the Ntn hydrolase superfamily (41). The observation that a pvdQ mutant of PAO1 retained its ability to grow on longchain AHLs suggested that other AHL-degrading enzymes were also employed by P. aeruginosa (42), and additional studies confirmed that two other Ntn hydrolase homologues, QuiP (PA1032) and HacB (PA0305), also function as AHL acylases with substrate specificity similar to that of PvdQ (45,52). Importantly, although PvdQ, QuiP, and HacB were all capable of modulating the accumulation of the endogenous P. aeruginosa signal 3OC12HSL (44,45,52), their abilities to modulate P. aeruginosa behavior and virulence in vivo were distinct.…”

“…To date, most bacteria known to be capable of AHL inactivation have been found to contain only one or two enzymes responsible for this activity. However, there are exceptions, including P. aeruginosa, which employs multiple AHL acylases (PvdQ, QuiP, HacB, and PA3922) (42,44,45,52). One unique exception is the bacterium Rhizobium sp.…”