Plants use pattern recognition receptors to defend themselves from microbial pathogens. These receptors recognize pathogen-associated molecular patterns (PAMPs) and activate signaling pathways that lead to immunity. In rice (Oryza sativa), the chitin elicitor binding protein (CEBiP) recognizes chitin oligosaccharides released from the cell walls of fungal pathogens. Here, we show that the rice blast fungus Magnaporthe oryzae overcomes this first line of plant defense by secreting an effector protein, Secreted LysM Protein1 (Slp1), during invasion of new rice cells. We demonstrate that Slp1 accumulates at the interface between the fungal cell wall and the rice plasma membrane, can bind to chitin, and is able to suppress chitin-induced plant immune responses, including generation of reactive oxygen species and plant defense gene expression. Furthermore, we show that Slp1 competes with CEBiP for binding of chitin oligosaccharides. Slp1 is required by M. oryzae for full virulence and exerts a significant effect on tissue invasion and disease lesion expansion. By contrast, gene silencing of CEBiP in rice allows M. oryzae to cause rice blast disease in the absence of Slp1. We propose that Slp1 sequesters chitin oligosaccharides to prevent PAMP-triggered immunity in rice, thereby facilitating rapid spread of the fungus within host tissue.

Plants have evolved efficient defense mechanisms known as priming and synergy, both of which can mobilize defense responses more extensively against successive pathogen invasion or simultaneous stimulation by different signal molecules. However, the mechanisms underlying these phenomena were largely unknown. In the present study, we used cultured rice cells and combination of purified MAMP molecules as a model system to study the mechanisms of these phenomena. We found that the pretreatment of rice cells with a low concentration of bacterial lipopolysaccharide (LPS) apparently primed the defense responses induced by successive N-acetylchitooctaose (GN8) treatment. On the other hand, simultaneous treatment with GN8 and LPS also resulted in the similar enhancement of defense responses observed for the LPS-induced priming, indicating that the synergistic effects of these MAMPs are basically responsible for such enhancement of defense responses, though the effect could be interpreted as “priming” under some experimental conditions. These results also indicate that such a positive crosstalk of signaling cascade downstream of MAMP receptors seems to occur very rapidly, probably at early step(s) of signaling pathway. Comprehensive analysis of phytohormones revealed a specific enhancement of the synthesis of jasmonic acid (JA), both in the LPS pretreatment and also simultaneous treatment, indicating a role of JA in the enhancement of downstream responses.

Chitinase treatment of a commercial laminarioligosaccharide preparation from a mushroom resulted in a loss of previously reported elicitor activity in rice cells, indicating that the activity was attributable not to the laminarioligosaccharide but rather to the contaminating chitin fragments. This suggests that the elicitor activity of laminarioligosacchraides from such sources containing chitinaceous substances should be carefully interpreted.Key words: microbe-associated molecular pattern (MAMP); -glucan; chitin; elicitor; rice Branched -glucans are a typical component of the cell walls of various fungi and oomycetes, and hence are recognized as a representative MAMP (microbeassociated molecular pattern) for these microbes.1,2) In fact, their fragments, -glucan oligosaccharides, have been found to act as potent elicitors in various plant species. A hepta--glucoside from Phytophthora megasperma f. sp. glycinea and a tetraglucosylglucitol from the rice blast fungus Pyricularia oryzae are two well characterized examples of elicitor-active -glucan oligosaccharides, acting specifically on certain legumes and rice, respectively.3,4) Both of these oligosaccharides are branched at the O-3 or the O-6 positions of glucosyl residues in their 1,6-linked or 1,3-linked backbone chains and the positions of branching are critical to their biological activities. On the other hand, several papers have reported that linear laminarioligosaccharides also serve as an elicitor in some plant cells, including rice. 5,6)As we did not find any elicitor activity for linear laminarioligosaccharides in rice, 4) we tried to clarify the reason for the discrepancy, and found that contamination by minute amounts of chitin oligosaccharides in some commercial laminarioligosaccharides prepared from a mushroom caused the difference, at least in one case. Figure 1 shows the elicitor activity of two commercial products of laminaripentaose prepared from pachyman, a 1,3-linked -glucan from the cell walls of a mushroom, or curdlan, also a 1,3-linked -glucan from a bacterium, on suspension-cultured rice cells. In both the generation of reactive oxygen species (ROS; Fig. 1, top) and the induction of a defense gene (Fig. 1, bottom), OsKSL4, encoding an enzyme involved in the biosynthesis of phytoalexin, 7,8) only the laminaripentaose preparation from pachyman showed extensive elicitor activity comparable to that of N-acetylchitooctaose (GN8). 9)Considering the reason for this difference in elicitor activity between these two products, we noticed that the one prepared from pachyman might contain a small amount of chitin oligosaccharides, because chitin is a typical cell wall component of fungal cell walls including mushrooms, whereas bacterial cell walls do not contain chitin. To examine this possibility, laminaripentaose from pachyman was treated with rice chitinase expressed in yeast 10) and purified by affinity chromatography, and analyzed for elicitor activity. As shown in Fig. 2, the elicitor activity shown by the limanaripentaose preparation disappea...