Suppression of Plant Defence Reactions in Alfalfa Cell Cultures by Sinorhizobium meliloti Surface Carbohydrates

Abstract

The soil bacterium Sinorhizobium meliloti is able to establish a symbiosis with its host plant alfalfa (Medicago sativa) as well as with the model legume Medicago truncatula. During the establishment of this symbiosis bacterial cells infect plant roots and induce a new organ, the root nodule. The bacterial nod genes, code for the synthesis and excretion of the nodulation factors (Nod factors), which are able to induce nodule organogenesis in the legume. The nodulation factors are structurally related to chitin, a common constituent of the fungal cell wall which is absent in higher plants. Chitin fragments are known to act as elicitors of defence reactions in cell cultures of various plant species. Because of the structural similarity between chitin elicitors and Nod factors we analysed whether these compounds may stimulate the plant defence system of M. sativa. We first established elicitor-responsive cell suspension cultures of Medicago sativa (host plant) and Nicotiana tabacum (non-host plant) as a system to test the perception of biological active compounds (Baier et al, in press). Alfalfa suspension cultures respond to yeast elicitors with a strong alkalinization of the culture medium and a transient synthesis of activated oxygen species, generally termed oxidative burst. The alkalinization reaction as well as the oxidative burst were also observed when tobacco (N. tabacum) cell suspension cultures were treated with yeast elicitors. Dependent on the degree of polymerisation N-acetyl chitin oligomers induced the alkalinization response in both plant cell suspension cultures, while only tobacco cell cultures developed an oxidative burst. Suspension-cultured tobacco cells responded to S. meliloti nodulation factors with an alkalinization of 0.25 pH units and a significant oxidative burst. In contrast, addition of S. meliloti nodulation factors to suspension-cultured alfalfa cells induced no oxidative burst. Instead of an alkalinization, they showed a slight acidification of the culture medium. Apparently, alfalfa might have evolved a perception system that discriminates between nodulation factors and chitin oligosaccharides. Beside the nodulation factors surface carbohydrates of S. meliloti play a curical role in the establishment of the symbiosis (Niehaus and Becker, 1998). Mutants of S. meliloti that failed to synthesize the acidic exopolysaccharide EPS I (succinoglycan) were unable to induce effective root nodules. In addition these strains induced a defence response on their host plants, probably preventing the infection by the symbiont (Niehaus et al., 1993). Further on, the bacterial lipopolysaccharide (LPS) plays a role in the interaction of S. meliloti with its different host plants. The specific S. meliloti LPS-mutant Rm6963 (Lagares et al. 1992), mutated in lpsB, was shown to induce effective nodules on M. sativa, but failed to establish an effective symbiosis with M. truncatula (Niehaus et al. 1998). M. truncatula root nodules induced by Rm6963 also exhibited symptoms of plant defence reactions. From these observations we propose that rhizobial surface carbohydrates act as suppressors of the plant defence system, enabling the symbiont S. meliloti to infect the host plant. Using the cell culture assay system, purified homologous and heterologous LPS were analysed for possible suppressor functions in alfalfa and tobacco plant suspension cultures. In alfalfa cell cultures the elicitor induced