Quorum sensing in nitrogen-fixing rhizobia

Abstract

Members of the rhizobia are distinguished for their ability to establish a nitrogen-fixing symbiosis with leguminous plants. While many details of this relationship remain a mystery, much effort has gone into elucidating the mechanisms governing bacterium-host recognition and the events leading to symbiosis. Several signal molecules, including plant-produced flavonoids and bacterially produced nodulation factors and exopolysaccharides, are known to function in the molecular conversation between the host and the symbiont. Work by several laboratories has shown that an additional mode of regulation, quorum sensing, intercedes in the signal exchange process and perhaps plays a major role in preparing and coordinating the nitrogen-fixing rhizobia during the establishment of the symbiosis. Rhizobium leguminosarum, for example, carries a multitiered quorum-sensing system that represents one of the most complex regulatory networks identified for this form of gene regulation. This review focuses on the recent stream of information regarding quorum sensing in the nitrogen-fixing rhizobia. Seminal work on the quorum-sensing systems of R. leguminosarum bv. viciae, R. etli, Rhizobium sp. strain NGR234, Sinorhizobium meliloti, and Bradyrhizobium japonicum is presented and discussed. The latest work shows that quorum sensing can be linked to various symbiotic phenomena including nodulation efficiency, symbiosome development, exopolysaccharide production, and nitrogen fixation, all of which are important for the establishment of a successful symbiosis. Many questions remain to be answered, but the knowledge obtained so far provides a firm foundation for future studies on the role of quorum-sensing mediated gene regulation in host-bacterium interactions.

FIG. 1.

Quorum-sensing model in P. fischeri. At low cell densities, transcription of the luxICDABE operon occurs at basal levels. LuxI encodes the AHL synthase, which synthesizes 3-oxo-C₆-HSL from acyl-ACP and SAM substrates. High cell densities lead to the accumulation of AHLs, which bind and activate the LuxR transcriptional activator. LuxR binds to an inverted repeat referred to as the lux box, which is centered at −42.5 from the transcriptional start site, and makes contact with the RNA polymerase to stimulate the expression of the luxICDABE genes. Expression of the luxR gene is regulated by several factors such as heat shock, catabolite repression, and even LuxR itself (only at very high cell densities).

FIG. 2.

Rhizobium-legume symbiosis model. The process of nodule invasion begins with the production, by the plant roots, of phenolic signals called flavonoids. Flavonoids serve as inducers for the NodD transcriptional activator, which then binds to conserved promoter elements, referred to as nod boxes, upstream of the nodulation genes (nod, noe, and nol). Expression of the nodulation genes results in the production of Nod factor molecules composed of a backbone of N-acetylglucosamine residues (two to five), a fatty acyl moiety with variable length and degrees of saturation, and various decorations on the backbone, all of which are species specific. Bacterial Nod factors then act on the plant roots to induce nodule formation and root hair curling. The bacteria are then able to invade the root hairs; this process requires the production of certain symbiotically active exopolysaccharides, which also vary depending on the species. Once inside, the bacteria undergo a differentiation process and begin expressing nitrogenase and other genes necessary for nitrogen fixation.

FIG. 3.

R. leguminosarum bv. viciae quorum-sensing network. R. leguminosarum harbors four known quorum-sensing systems. The cinRI system resides on the chromosome and produces 3-OH-C_14:1-HSL, which positively influences the tra and rai systems. BisR plays a dual role in activating traR and repressing cinR in response to 3-OH-C_14:1-HSL, thereby linking the cin and tra systems. pRL1JI harbors both the tra and rhi systems, as well as the genes that confer growth sensitivity in response to 3-OH-C_14:1-HSL. The tra system is responsible for the production of 3-oxo-C₈-HSL and controls conjugal plasmid transfer, while the rhi system produces several short-chain AHLs and influences nodulation efficiency by an unknown mechanism. The raiRI locus resides on pIJ9001 and also produces several short-chain AHLs; however, little is known about the role of this quorum-sensing system.

FIG. 4.

Interaction of the cinRI and raiRI quorum-sensing systems in R. etli CNPAF512. R. etli CNPAF512 carries two quorum-sensing systems, cinRI and raiRI, both of which are located on the chromosome. The cinRI locus encodes a long-chain AHL and is involved in growth inhibition, as in R. leguminosarum. This locus is also required for efficient nitrogen fixation and proper symbiosome development. The raiRI system is positively regulated by cinI-produced AHL and is, in turn, responsible for the production of several short-chain AHLs. The raiRI locus also controls nitrogen fixation and mediates growth inhibition.

FIG. 5.

Quorum sensing in R. etli CFN42. Quorum sensing in R. etli CFN42 controls conjugal plasmid transfer through the action of the tra genes, which are located on p42a. In this system, traI produces 3-oxo-C₈-HSL and is regulated by both CinR and TraR. The traM gene is present but does not seem to be expressed and therefore does not play a regulatory role in strain CFN42. A second AHL has also been described, but the locus responsible for its production has not been identified.

FIG. 6.

Quorum sensing in Rhizobium sp. strain NGR234. NGR234 carries a large symbiotic plasmid, pNGR234a, which harbors the tra quorum-sensing system. The tra genes control the production of 3-oxo-C₈-HSL as well as the conjugal transfer of pNGR234a. At least one other quorum-sensing system may be present elsewhere in the genome and controls the production of 3-oxo-C₈-HSL and another long-chain AHL.

FIG. 7.

S. meliloti quorum-sensing systems. S. meliloti strain Rm41 harbors three quorum-sensing systems, while strain Rm1021 carries only two systems. The tra system, which is carried on pRme41a and is present only in strain Rm41, produces several short-chain AHLs, including 3-oxo-C₈-HSL, and controls the conjugal transfer of pRme41a. The sinRI system resides on the chromosome and is present in both strains. SinI produces several long-chain AHLs, and at least one of them (C_16:1-HSL), along with ExpR, is required for the production of a symbiotically important exopolysaccharide, EPS II. The mel system, also present in both strains, produces several short-chain AHLs, but the corresponding genes remain to be identified.

FIG. 8.

Quorum sensing in B. japonicum. Quorum sensing in B. japonicum is involved in repressing the nodulation genes at high cell densities. The system, unlike other rhizobial quorum-sensing systems, is not controlled by AHLs. Instead, the autoinducer is a low-molecular-weight compound termed bradyoxetin. NwsB is part of a two-component system and is required for the detection of bradyoxetin. NwsB then induces nolA, which in turn induces nodD2, leading to repression of the nod genes.