Abstract
BackgroundThe protein encoded by GmRLK18-1 (Glyma_18_02680 on chromosome 18) was a receptor like kinase (RLK) encoded within the soybean (Glycine max L. Merr.) Rhg1/Rfs2 locus. The locus underlies resistance to the soybean cyst nematode (SCN) Heterodera glycines (I.) and causal agent of sudden death syndrome (SDS) Fusarium virguliforme (Aoki). Previously the leucine rich repeat (LRR) domain was expressed in Escherichia coli.ResultsThe aims here were to evaluate the LRRs ability to; homo-dimerize; bind larger proteins; and bind to small peptides. Western analysis suggested homo-dimers could form after protein extraction from roots. The purified LRR domain, from residue 131–485, was seen to form a mixture of monomers and homo-dimers in vitro. Cross-linking experiments in vitro showed the H274N region was close (<11.1 A) to the highly conserved cysteine residue C196 on the second homo-dimer subunit. Binding constants of 20–142 nM for peptides found in plant and nematode secretions were found. Effects on plant phenotypes including wilting, stem bending and resistance to infection by SCN were observed when roots were treated with 50 pM of the peptides. Far-Western analyses followed by MS showed methionine synthase and cyclophilin bound strongly to the LRR domain. A second LRR from GmRLK08-1 (Glyma_08_g11350) did not show these strong interactions.ConclusionsThe LRR domain of the GmRLK18-1 protein formed both a monomer and a homo-dimer. The LRR domain bound avidly to 4 different CLE peptides, a cyclophilin and a methionine synthase. The CLE peptides GmTGIF, GmCLE34, GmCLE3 and HgCLE were previously reported to be involved in root growth inhibition but here GmTGIF and HgCLE were shown to alter stem morphology and resistance to SCN. One of several models from homology and ab-initio modeling was partially validated by cross-linking. The effect of the 3 amino acid replacements present among RLK allotypes, A87V, Q115K and H274N were predicted to alter domain stability and function. Therefore, the LRR domain of GmRLK18-1 might underlie both root development and disease resistance in soybean and provide an avenue to develop new variants and ligands that might promote reduced losses to SCN.
Highlights
The protein encoded by GmRLK18-1 (Glyma_18_02680 on chromosome 18) was a receptor like kinase (RLK) encoded within the soybean
This report describes a functional analysis and structural prediction of the leucine rich repeat (LRR) domain from a RLK protein involved in plant pest and disease resistance
The results showed that the isolated LRR of GmRLK18-1 binds peptides found in both nematode and plant secretions with high affinity
Summary
The protein encoded by GmRLK18-1 (Glyma_18_02680 on chromosome 18) was a receptor like kinase (RLK) encoded within the soybean The locus underlies resistance to the soybean cyst nematode (SCN) Heterodera glycines (I.) and causal agent of sudden death syndrome (SDS) Fusarium virguliforme (Aoki). The leucine rich repeat (LRR) domain was expressed in Escherichia coli Plants employ both cell surface and cytoplasmic receptors to respond to a wide array of signals from pathogens [1]. Merr.) are the soybean cyst nematode (SCN; Heterodera glycines I.) and sudden death syndrome (SDS) agent Fusarium virguliforme (Aoki) [3]. Rhg on chromosome 8 (linkage group (Lg) A2) and Rhg1/Rfs on chromosome 18 (Lg G), contain genes that encode receptor like kinase (RLK) proteins within the RPK gene family implicated in resistance. Only GmRLK18-1 has been shown to underlie resistance to both pathogens in transgenic plants [8,9]
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