## 2 Quorum Sensing Genes in _Sinorhizobium melilioti_  
Most of the knowledge of QS in _S. melilioti_ has been obtained using the reference strain Rm1021. In this bacterium, three genes, \(sinl,sinR\), and \(expR\), are essential for QS regulation (Figure 1). _sinl_ encodes a Luxl-type synthase that catalyzes AHL production from two substrates: _S_-adenosyl-L-methionine (SAM), which provides the homoserine lactone ring moiety, and an appropriate acyl-acyl carrier protein (acyl-ACP) from fatty acid biosynthesis, which is the precursor of the fatty acyl side chain. SinI is responsible for the production of diverse long-chain AHLs, including C12-HL, C14-HL, 3-oxo-C14-HL, C16-HL, 3-oxo-C16-HL, C16-HL, 3-oxo-C16:1-HL and C18-HL, with 3-oxo-C16:1-HL being the most abundant in laboratory cultures [24; 25; 26]. Adjacent and upstream of the chromosomal \(sinl\) gene is \(sinR\), which encodes a LuxR-type transcription regulator that controls \(sinl\) expression but whose activity is not affected by AHLs [27; 28; 24]. Instead, the product of the \(expR\) gene is the major regulator of AHL-controlled gene expression in _S. melilioti_[26; 29; 30]. ExpR is an orphan LuxR-type transcriptional regulator or \"LuxR solo\", i.e. a LuxR-type receptor that is not paired with a cognate AHL synthase [31; 32]. Expression of \(sinl\) is strongly enhanced by ExpR in the presence of AHLs, resulting in a positive feedback. This positive feedback mechanism is a hallmark feature of the classical QS system [33]. As will be explained in detail in Section 3.1.1 of this review, ExpR also represses transcription of \(sinR\) at high AHL concentrations, which leads to negative feedback regulation of \(sinl\)[34] (Figure 1). Compared to other orphan LuxR-type proteins, ExpR is a highly versatile regulator with different regulatory capabilities. More than 500 genes have been identified as differentially expressed by ExpR [30] and over 30 DNA binding sites have been biochemically identified [28; 35]. Although ExpR can act as an AHL-independent regulator of genes involved in metabolic processes, the presence of AHLs is necessary for most of the regulatory targets of ExpR, including the two best-known functions controlled by QS in _S. melilioti_: activation of EPS synthesis and repression of flagellaproduction (see Section 5; Figure 1). The location of the ExpR-binding site within the promoter region, e.g., upstream or downstream of the \(-35/-10\) promoter elements, determines whether ExpR/AHL activates or represses transcription. In addition, transcriptional activity of QS-regulated genes is dependent upon the concentration of AHLs [28]. This property allows for a temporal expression program in which different target promoters have different AHL concentration thresholds for their activation or repression. As an example, genes required for EPS production in _S. melidi_ are activated at lower concentrations of AHLs than those required for the repression of flagella-associated genes [28; 30]. The _expR_ gene is disrupted by an insertion element in the widely studied _S. melidi_ laboratory strains Rm1021 and Rm2011 and its functional restoration causes a moderate (3-fold) increase in production of the symbiotically important EPS succinoglycan (EPS I) and an especially strong increase in galactoglucan (EPS II) production, which confers an extremely mucoid phenotype to the colonies [36; 37; 38; 39]. Remarkably, the dry phenotype exhibited by various laboratory strains of _S. melidi_ from multiple independent isolates has been accredited to spontaneous mutations at the _expR_ locus [36; 40]. It was found that _expR_ and therefore QS is rapidly inactivated during routine laboratory  
Figure 1: Model of transcriptional and post-transcriptional regulation of quorum sensing (QS) in _Sinorhizobium melidi_. Transcriptional autoregulation of the ExpR/Sin system is shown. _N_-acyl-homoserine lactone (AHL)-activated ExpR induces _sinl_ and inhibits _sinR_. AHL-activated ExpR activates _expR_ expression weakly.