Rhizobium Symbiosis: Insight into Nod Factor Receptors

The symbiotic association between Rhizobia and legume plants leads to the formation of nitrogenfixing nodules, conferring on host plants the ability to be grown without the addition of nitrogen fertiliser. This symbiosis is characterised by a high level of host specificity, mediated by specific recognition of rhizobial molecules called Nod factors [1]. These are lipochito-oligosaccharides, based on a backbone of generally four or five N-acetyl glucosamine residues which are N-acylated at the non-reducing end and carry various substitutions. The nature of the N-acyl chains and the other substitutions are important determinants of rhizobial host specificity. The infection process that leads to the Rhizobium– legume symbiosis is controlled by the Nod factors; Rhizobia need them to be able to induce specialised structures called infection threads, through which they penetrate into the cortex of host roots. Moreover, purified Nod factors are active in roots of host plants and, at pico/nanomolar concentrations, they can induce many of the responses characteristic of the bacteria themselves. These include both rapid, localised responses, such as calcium fluxes in root hair cells, and slower ones that encompass several cell layers of the root, for example the induction of gene expression and cell division that leads to the formation of nodule primordia. Nod factors thus act as symbiotic signalling molecules for conferring host specificity, for infection and for nodule development, and are likely to be perceived by high affinity receptors [2]. The recent cloning of legume genes has revealed a new class of receptors containing LysM domains which function in Nod factor perception [3–5]. Genetic dissection of the mechanisms by which the Nod factor signal is perceived and transduced by host plants led to the identification of genes that control different steps of a Nod factor-activated signalling pathway [6–8]. Some of these genes control both early steps of Nod factor signalling and the establishment of another, more ancient type of root endosymbiosis — the arbuscular mycorrhizal symbiosis. Such ‘common symbiotic’ genes define what is probably an ancient signalling pathway that has been recruited by legumes and Rhizobia and might be involved in transducing both fungal and rhizobial signals [6,9]. Genes which are good candidates to code for Nod factor receptors have been identified on the basis that they intervene only in the Rhizobium–legume symbiosis, and because alterations in them result in plants that either no longer respond to Nod factors or that show an altered specificity directly related to Nod factor structure. By exploiting the model legumes Medicago truncatula and Lotus japonicus, two genes in each of these two categories have recently been cloned [3–5], and all four genes are predicted to code for transmembrane serine/threonine receptor-like kinases containing extracellular LysM domains (LysM-RLKs). The genes NFR1 and NFR5, isolated by positional cloning in L. japonicus, are required for the earliest physiological and cellular responses to Nod factors, as mutations in them lead to plants that either no longer respond to Nod factors or show attenuated responses [3,4]. NFR1 and NFR5, like most genes that control Nod factor signalling, also control root hair curling, the first step of rhizobial infection, preceding infection thread formation [3,4]. The two other cloned genes, LYK3 and LYK4, were identified in M. truncatula as candidate Nod factor receptor genes in two steps [5]. First, they were found to occur in a region of the M. truncatula genome syntenic to the region of the pea genome containing the gene SYM2; a specific allele of this gene from Afghanistan pea, SYM2A, confers a high demand on Nod factor structure with respect to nodulation and infection thread formation. Second, reverse genetics and rhizobial strains producing different Nod factor structures were used to study whether knock-down lines of M. truncatula differed from wild-type plants in their Nod factor structural dependency for nodulation and infection thread formation. Both phenotypes were affected for LYK3 knock-downs, while LYK4 knockdowns just showed aberrant infection thread morphology [5]. As infection of the knock-down plants is essentially arrested at the stage of entrapment in curled root hairs with only abnormal infection threads forming, in the presence of rhizobia producing modified Nod factors, it is proposed that the LYK3 and LYK4 proteins control bacterial entry into root hairs and infection thread formation in a manner dependent on Nod factor structure. NFR1, NFR5, LYK3 and LYK4 are all predicted to contain a signal peptide, extracellular LysM domains, a transmembrane segment and an intracellular serine/threonine kinase domain [3–5] (Figure 1). Several features of these proteins indicate that two classes can be distinguished, one consisting of NFR1, LYK3 and LYK4, which all have two LysM domains, and the other with NFR5, which has three LysM domains [3–5]. The genes encoding NFR1, LYK3 and LYK4 all have 12 exons in conserved positions, while NFR5 is encoded by a single exon. There is a high level of sequence Dispatch Current Biology, Vol. 13, R973–R975, December 16, 2003, ©2003 Elsevier Science Ltd. All rights reserved. DOI 10.1016/j.cub.2003.11.047