Monitoring symbiotic gene expression in Rhizobium sp. NGR234

Rhizobium sp. NGR234 has the largest capacity to nodulate legumes of any soil bacterium known. Its genome comprises three replicons: a 536 kb symbiotic plasmid, a c.a. 2,000 kb mega-plasmid, and a 3,700 kb chromosome. Sequencing the 536 kb plasmid, pNGR234a showed that apart from nodE, nodG, and nodPQ, most Nod-factor biosynthetic genes are plasmid-borne, and regulated by a number of transcriptional regulators of the LysR family. Of these, NodD1 is of primary importance. It is activated by a variety of phenolic compounds found in the rhizospheres of plants, and the NodD1-flavonoid complex enhances expression of nod-box controlled genes. A second part of the broad host-range of NGR234 can be attributed to the 16 nod-genes which are responsible for the elaboration of a large family (>80 members) of lipo-oligosaccharidic Nod-factors. A further 10 genes (rhcC1 to rhcV) encode functions necessary for the elaboration of the bacterial type III protein secretion system (TTSS). The TTSS is an essential component of pathogenicity in both animal (e.g. Salmonella, Shigella, Yersinia) and plant (Erwinia, Pseudomonas, Ralstonia, and Xanthomonas) pathogens. Secretion of at least two proteins is controlled by the TTSS of NGR234 and occurs after flavonoid induction. Inactivation of the TTSS by insertional mutagenesis blocks the secretion of both proteins, and strongly affects the ability of NGR234 to nodulate a variety of tropical legumes including Pachyrhizus tuberosus and Tephrosia vogelii. High resolution transcription analysis of pNGR234a shows that many more genes than the known nod, nol, rhc, nif and fix loci are transcribed under symbiotic conditions. This, together with the large number of newly discovered symbiotic promoters suggests that many more genes than previously thought are necessary for a functional symbiosis.