The regulatory locus cinRI in Rhizobium leguminosarum controls a network of quorum-sensing loci

Abstract

N-(3-hydroxy-7-cis-tetradecenoyl)-L-homoserine lactone (3OH,C(14:1)-HSL) is a quorum-sensing signalling molecule produced by Rhizobium leguminosarum. It is unusual in that it inhibits the growth of several strains of R. leguminosarum and was previously known as 'small bacteriocin'. The cinRI locus responsible for the production of 3OH,C(14:1)-HSL has been characterized; it is predicted to be on the chromosome, based on DNA hybridization. The cinR and cinI genes are in different transcriptional units, separated by a predicted transcription terminator. CinR regulates cinI expression to a very high level in a cell-density dependent manner, and cinI expression is positively autoregulated by 3OH,C(14:1)-HSL, the only identified N-acyl homoserine lactone (AHL) produced by CinI. No other AHLs were identified that strongly induced cinI expression. Mutation of cinI or cinR abolishes the production of 3OH,C(14:1)-HSL and also reduces the production of several other AHLs. This is thought to result from the expression of three other AHL production loci being affected by the absence of 3OH,C(14:1)-HSL. AHLs produced by these other loci include N-hexanoyl- and N-octanoyl-L-homoserine lactones and, unexpectedly, N-heptanoyl-L-homoserine lactone (C7-HSL). The expression of the rhiI gene on the symbiotic plasmid is greatly reduced in a cinI mutant, and the major regulatory effect appears to be mediated at least in part as a result of an effect on expression of RhiR, the regulator of rhiI. Thus, cinR and cinI appear to be at the top of a regulatory cascade or network that influences several AHL-regulated quorum-sensing loci. The expression of cinI-lacZ fusions is significantly reduced (but not abolished) when the symbiosis plasmid pRL1JI is present, resulting in a reduction in the level of 3OH,C(14:1)-HSL produced. Mutation of cinI had little effect on growth or nodulation. However, plasmid transfer was affected, and the results obtained indicate that 3OH,C(14:1)-HSL produced by either the donor or the recipient in mating experiments can stimulate transfer of pRL1JI.