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From 2011.igem.org

Modern perspective

        Actually, quorum sensing system (QS) is highly concentrated on in microbiology field. Quorum sensing is a system of stimulus and response correlated to population density. Many species of bacteria use quorum sensing to coordinate gene expression according to the density of their local population. Quorum sensing can function as a decision-making process in any decentralized system, as long as individual components have: (a) a means of assessing the number of other components they interact with and (b) a standard response once a threshold number of components is detected (by Wikipedia).
        “QS” is thought to be the “language” among bacteria. By using it, bacteria can make some “social” behaviors. Such as Vibrio harveyi in ocean, they can get together and light up a piece of water.
        “QS” can also use as a switch. As we do in our project, AHL signal molecule is produced by one kind of bacteria’s QS system. And another one receives it to promote under genes. When the quality of first bacteria gets to a setting number, it can begin to “wake up” the second one. And thus can make a cycle.
        In the following, we will make a detailed introduction of the quorum-sensing system.

Quorum-Sensing System Introduction

        Bacterial populations coordinately regulate gene expression by producing diffusible signal molecules. These signals, known as autoinducers, accumulate extracellularly and interact specifically with a receptor protein to affect changes not related to their own metabolism. Production of autoinducers typically occurs at specific stages of growth or in response to changes in the environment and induces a concerted response once a critical concentration has been reached. These diffusible signals frequently act to induce gene expression in response to bacterial cell density in a process often referred to as quorum sensing. Alternatively, autoinducer secretion and response may confer on the bacterium the ability to determine whether secreted molecules move away from the cell. This process, termed diffusion sensing by Rosemary Redfield, could allow the cells to regulate the secretion of effectors, such as degradative enzymes, antibiotics, surfactants, and siderophores, to minimize losses to extracellular diffusion. The best characterized quorum-sensing mechanism is found in gram-negative organisms and involves the use of acylated homoserine lactones (AHLs) as signal molecules.

HOW BACTERIA TALK TO EACH OTHER？

AHL-Mediated Cell-Cell Communication
        Historically, it was thought that bacteria were solitary individuals, each growing independently of the population. However, in 1970 Nealson et al. discovered that bacteria can sense and respond to the rest of the population. This phenomenon is called quorum sensing and is defined as the cell density-dependent regulation of gene expression. One of the best-studied examples of quorum sensing is in Photobacterium fischeri (formerly Vibrio fischeri), a marine bacterium that is a symbiont of several marine fish and squids (Fig. 1) . In this model organism, the basal-level synthesis of autoinducers occurs at low cell densities, like those found in seawater. The autoinducers, which belong to the AHL family of signal molecules, are thought to pass through the cell membrane by diffusion . As the cell density increases during the symbiotic association with the animal host, autoinducers accumulate in and around the cells .
        When a threshold level of AHLs (about 10 nM) is reached, the LuxR regulator is activated by binding the AHL .LuxR, a transcriptional activator, then induces expression of the lux operon (Fig. 1). The lux operon contains luxI (the AHL synthase) along with the genes necessary for luminescence. Activation of the lux operon leads to a rapid rise in the levels of autoinducer and creates a positive-feedback loop, which is followed by the onset of luminescence. LuxR is regulated at the transcriptional level by cyclic AMP receptor protein and presumably at the posttranscriptional level by GroEL . At low AHL levels, LuxR activates its         expression, while at high AHL levels, the active LuxR represses itself.

        Next，we want to have a brief introduction of the molecular mechanisms of AHL family QS system.

MOLECULAR MECHANISMS

AHL Family of Autoinducers
        AHL characteristics. AHLs consist of an HSL head group attached to a variable acyl side chain (as below). The amphipathy of the AHL molecule seems to be a balance between the hydrophobic side chain and the hydrophilic HSL ring. These characteristics presumably allow the AHLs to traverse the phospholipid bilayer of the cell membrane and to navigate the aqueous intracellular and extracellular environments . The acyl chain varies in length, from 4 to 18 carbons in those AHLs identified so far. Variability also exists in the third carbon position of the acyl chain, where there can be a hydrogen, hydroxyl, or oxo substitution (as below). A few AHLs that have unsaturated acyl chains have also been identified. The overall length of the side chain and the chemical modification at the third carbon position provide the specificity to quorum-sensing signals. To add complexity,most organisms produce more than one type of AHL and different organisms can produce the same AHL. Therefore, there is some overlap in the production and recognition of AHLs by different organisms.

        LuxI-type synthases. All LuxI-type proteins identified to date resemble the LuxI of P. fischeri and catalyze the ligation of S-adenosylmethionine (SAM) with an acylated acyl carrier protein (acyl-ACP), which form the HSL and acyl chain components, respectively, of the resulting AHL. The catalytic model proposed by Parsek, Greenberg, and coworkers suggests a nucleophilic attack on the C-1 position of the acyl-ACP molecule by the amino nitrogen of SAM, resulting in an amide linkage. This is followed by the lactonization of SAM together with amide bond formation, which results in ring formation and release of the AHL. LuxI homologues are about 200 amino acids in length, and disruption of certain conserved residues in the amino-terminal half of the protein leads to a reduction or loss of synthase activity. It has been suggested that conserved amino acids in the carboxy
        terminus may be necessary for acyl-ACP selection. The recently determined crystal structure of EsaI, the LuxI homologue from P. stewartii, shows remarkable similarity to N-acetyltransferases, and its core catalytic fold has features essential for phosphopantetheine binding. The structural analysis suggests that the N acylation of SAM is likely to include abstraction of an amine proton by a catalytic base. In addition, variable residues in the C-terminal half of the protein and the nature of the amino acid at position 140 constitute the basis for the acyl chain specificity.
        Besides，there exists LuxM/AinS-type synthases and LuxS-type synthases. For more information， please see the full text of Quorum Sensing in Nitrogen-fixing Rhizobia on the bottom of this page.

Quorum-Sensing Regulators
        LuxR-type regulators. LuxR-type proteins share two regions of sequence conservation, an AHL binding domain and a DNAbinding motif. The current model suggests thatLuxR acts as a dimeric protein. LuxR regulators have an amino-terminal domain that binds AHLs and mediates protein oligomerization. The cytoplasmic carboxy-terminal domain includes a helix-turn-helix DNA binding region that is thought to be involved in transcriptional regulation. Specific interactions between the cognate AHLs and purified LuxR homologues have been demonstrated by various laboratories. Studies of A. tumefaciens have suggested that TraR, on binding the AHL signal, undergoes a conformational change, dimerizes, and activates transcription. Transcriptional activation by LuxR-type proteins requires cis-acting DNA elements, normally referred to as lux-typeboxes. The typical lux-type box is an 18- to 22-bp inverted-repeat sequence centered at about -40 from the transcriptional start site. Although many target genes of the LuxR-type regulators contain lux-type boxes within their promoters, there are reports of targets lacking a discernible lux box. Once bound, LuxR facilitates the binding of RNA polymerase to the target promoter, leading to activation of transcription. Although most instances show AHLbound LuxR-type proteins to function as transcriptional activators, a few examples of these proteins have been reported to function as repressors.

AHL Accumulation and Transport
        Although AHLs can clearly accumulate as a result of a simple increase in bacterial numbers, other factors could also influence the environmental concentration of these signal molecules. Bacterial aggregation, biofilm formation, and physical confinement could play roles in increasing the local concentration of AHLs. Other conditions such as high pH or enzymatic degradation could effectively decrease the concentration of signal molecules available for LuxR-type protein activation.
        All of the information above is from the Quorum Sensing in Nitrogen-fixing Rhizobia by Juan E.
Gonza´lez and Melanie M. Marketon. For more information, see the full text.