A BSTR ACTRhizobial lipo-chitooligosaccharides (LCOs) are signaling molecules involved in host-range recognition for the establishment of the symbiosis with leguminous plants. The major LCO of Rhizobium meliloti, the symbiont of Medicago plants contains four or five N-acetylglucosamines, O-acetylated and N-acylated with a C16:2 fatty acid on the terminal nonreducing sugar and O-sulfated on the reducing sugar. In this paper, the ligand specificity of a high-affinity binding site (Nod factor binding site 2 or NFBS2), enriched in a plasma membrane-enriched fraction of Medicago cell suspension cultures, is reported. By using chemically synthesized LCOs, the role of structural elements, important for symbiotic activities, as recognition motifs for NFBS2 was determined. The results show that the substitutions on the nonreducing sugar of the LCOs (the O-acetate group, the fatty acid, and the hydroxyl group on the C 4 of the sugar) are determinants for high-affinity binding to NFBS2. In contrast, the sulfate group, which is necessary for all biological activities on Medicago, is not discriminated by NFBS2. However, the reducing sugar of the LCO seems to interact with NFBS2, because ligand binding is affected by the reduction of the free anomeric carbon and depends on the number of N-acetyl glucosamine residues. These results suggest that the recognition of the LCOs by NFBS2 is mediated by structural elements in both the lipid and oligosaccharidic moities, but not by the sulfate group.Rhizobia are bacteria that can elicit the formation of nodules on the roots of legumes in which they fix dinitrogen for the benefit of the plant. Nodule formation is very specific, and a given strain of Rhizobium can infect a limited number of species of the legume family. The establishment of the symbiosis depends on a particular class of signaling molecules produced by the prokaryotic partner: the Nod factors. Nod factors are lipo-chitooligosaccharides (LCOs) consisting of a N-acetylglucosamine backbone, N-acylated on the terminal nonreducing sugar. Variations in the number of glucosamine residues (three to five), in the structure of the fatty acid chain, and the presence of different substitutions on the oligosaccharidic backbone are characteristics of each bacterial strain. At subnanomolar concentrations, purified Nod factors provoke the early symbiotic responses in the plant and in certain species they can induce the morphogenesis of bacteria-free nodules (reviewed in refs.

The lipo-chitooligosaccharidic Nod factors produced by rhizobia are key molecules in the establishment of symbiosis with legumes and probably are recognized by the host plant via specific receptors. Here, we report on the presence of a binding site in cell cultures of Phaseolus vulgaris displaying a high affinity for Nod factors from Rhizobium tropici (NodRt-V) (Me, S, C18:1), a symbiont of this legume. The binding site shares common properties with NFBS2, a Nod-factor binding site previously characterised in Medicago varia, in terms of affinity, preferential plasma-membrane location, and sensitivity to proteases and lysine reactive reagents. However, the bean site poorly recognizes the Nod factors produced by Sinorhizobium meliloti, the symbiont of Medicago. The study of selectivity toward the Nod factors reveals that the length and degree of unsaturation of the acyl chain and the length of the oligosaccharidic moiety are important determinants of high affinity binding to the bean site; whereas, the N-methyl and O-sulfuryl groups play a minor role. Thus, the common characteristics of P. vulgaris and M. varia Nod-factor binding sites suggest that they probably correspond to structurally related proteins, but their different selectivity suggests that they may be involved in a differential perception system for Nod factors in legumes.

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