Final Activity Report Summary - QUORMETAB (Metabolic consequences of quorum sensing in bacteria)
For many years bacteria were considered only as autonomous single-celled organisms with little capacity for collective behaviour. However, we now appreciate that bacterial cells are in fact highly communicative and possess an extraordinary capacity for social behaviour. Bacteria coordinate their activities by producing and detecting small diffusible signal molecules which allow a population of bacteria to act as a group rather than as individuals. Such a cooperative behaviour is known as quorum sensing (QS) and plays a pivotal role in the lifestyle of both beneficial and pathogenic bacteria. Consequently, by increasing our understanding of how QS systems work, we can exploit bacterial cell-to-cell communication in order to discover better ways of controlling disease-causing bacteria and improve industrial and agricultural use of beneficial micro-organisms.
A significant amount of knowledge was accumulated on the chemical nature of QS signalling molecules (QSSMs) as well as on the molecular basis by which QS systems operate. However, despite the knowledge that production of QSSM was directly linked to the availability of starting molecules generated through the assimilation of nutrients by the cell, there were no studies determining the burden that QSSM production had upon metabolism or related to changes in metabolism affect QSSM production. Consequently, there was no published information on the metabolic control of QSSM synthesis as a function of growth environment, growth rate or population density or on the metabolic burden imposed by the presence of one or more QS circuits on a given organism.
Using pseudomonas aeruginosa as a model organism, our specific aims were to determine:
1. how growth phase, growth rate, nutrient limitation and cell population density influenced QS signal molecule (QSSM) synthesis and biosynthesis through the modulation of metabolism
2. how the manipulation of the activated methyl cycle (AMC) by mutagenesis of AMC pathway enzymes impacted on QS
3. what was the nature of the metabolic burden imposed by QSSM biosynthesis
4. how mutations in an efflux pump resulted in the loss of QSSM production through altered metabolism.
Our results demonstrated that, by exploiting quorum sensing signalling molecules including N-acyl-homoserine lactones (AHLs) and 2-alkyl-4-quinolones (AQs), which were derived from central precursors of the AMC such as S-adenosylmethionine (SAM), bacteria controlled their collective behaviour at the metabolic and genetic level. Firstly, we showed that QS was regulated by growth rate and cell population density. Particularly, under high population density and rapid growth, there was a considerable LasR-independent synthesis, i.e. the LasI and LasR QS system was placed on top of the hierarchy in the QS circuit, of the AQ signalling molecules. Hence, p. aeruginosa was adaptable at synthesising AQs. This could be especially important in environments such as cystic fibrosis lung where bacteria reproduced at high population density and maximum growth rate and where lasR mutants were often isolated and AQs were still isolated. Secondly, we demonstrated that QS had an important impact on the AMC, which provided precursors for both QS signalling molecules synthesis and a number of vital physiological conversions, and therefore imposed a metabolic burden on bacteria. Thirdly, we discovered that the MexGHI-OmpD efflux pump, which was previously shown to have a strong negative effect on QS and especially on AQs synthesis and was hypothesised to take part in the transport of AQs and their precursors, was involved in a more complex turnover of AQ biosynthesis than we expected, depending on the cell population density and the bacterial growth rate.
A significant amount of knowledge was accumulated on the chemical nature of QS signalling molecules (QSSMs) as well as on the molecular basis by which QS systems operate. However, despite the knowledge that production of QSSM was directly linked to the availability of starting molecules generated through the assimilation of nutrients by the cell, there were no studies determining the burden that QSSM production had upon metabolism or related to changes in metabolism affect QSSM production. Consequently, there was no published information on the metabolic control of QSSM synthesis as a function of growth environment, growth rate or population density or on the metabolic burden imposed by the presence of one or more QS circuits on a given organism.
Using pseudomonas aeruginosa as a model organism, our specific aims were to determine:
1. how growth phase, growth rate, nutrient limitation and cell population density influenced QS signal molecule (QSSM) synthesis and biosynthesis through the modulation of metabolism
2. how the manipulation of the activated methyl cycle (AMC) by mutagenesis of AMC pathway enzymes impacted on QS
3. what was the nature of the metabolic burden imposed by QSSM biosynthesis
4. how mutations in an efflux pump resulted in the loss of QSSM production through altered metabolism.
Our results demonstrated that, by exploiting quorum sensing signalling molecules including N-acyl-homoserine lactones (AHLs) and 2-alkyl-4-quinolones (AQs), which were derived from central precursors of the AMC such as S-adenosylmethionine (SAM), bacteria controlled their collective behaviour at the metabolic and genetic level. Firstly, we showed that QS was regulated by growth rate and cell population density. Particularly, under high population density and rapid growth, there was a considerable LasR-independent synthesis, i.e. the LasI and LasR QS system was placed on top of the hierarchy in the QS circuit, of the AQ signalling molecules. Hence, p. aeruginosa was adaptable at synthesising AQs. This could be especially important in environments such as cystic fibrosis lung where bacteria reproduced at high population density and maximum growth rate and where lasR mutants were often isolated and AQs were still isolated. Secondly, we demonstrated that QS had an important impact on the AMC, which provided precursors for both QS signalling molecules synthesis and a number of vital physiological conversions, and therefore imposed a metabolic burden on bacteria. Thirdly, we discovered that the MexGHI-OmpD efflux pump, which was previously shown to have a strong negative effect on QS and especially on AQs synthesis and was hypothesised to take part in the transport of AQs and their precursors, was involved in a more complex turnover of AQ biosynthesis than we expected, depending on the cell population density and the bacterial growth rate.