An integrated approach reveals how lipo‐chitooligosaccharides interact with the lysin motif receptor‐like kinase MtLYR3

Abstract

N-acetylglucosamine containing compounds acting as pathogenic or symbiotic signals are perceived by plant-specific Lysin Motif Receptor-Like Kinases (LysM-RLKs). The molecular mechanisms of this perception are not fully understood, notably those of lipo-chitooligosaccharides (LCOs) produced during root endosymbioses with nitrogen-fixing bacteria or arbuscular mycorrhizal fungi. In Medicago truncatula, we previously identified the LysM-RLK LYR3 (MtLYR3) as a specific LCO-binding protein. We also showed that the absence of LCO binding to LYR3 of the non-mycorrhizal Lupinus angustifolius, (LanLYR3), was related to LysM3, which differs from that of MtLYR3 by several amino acids and, particularly, by a critical tyrosine residue absent in LanLYR3. Here, we aimed to define the LCO binding site of MtLYR3 by using molecular modelling and simulation approaches, combined with site-directed mutagenesis and LCO binding experiments. 3D models of MtLYR3 and LanLYR3 ectodomains were built, and homology modelling and molecular dynamics (MD) simulations were performed. Molecular docking and MD simulation on the LysM3 identified potential key residues for LCO binding. We highlighted by steered MD simulations that in addition to the critical tyrosine, two other residues were important for LCO binding in MtLYR3. Substitution of these residues in LanLYR3-LysM3 by

Evolution of Lipochitooligosaccharide Binding to a LysM-RLK for Nodulation in Medicago truncatula

“…This hypothesis is supported by the AM phenotype of Mtlyk9 mutants (LYK-I subclass; Gibelin-Viala et al 2019), which is enhanced in a Mtlyk9/Mtnfp double mutant (Feng et al 2019). Other contenders for a role in AM in M. truncatula are MtLYR3 (LYR-IIIA subclass) since loss of LCO binding in MtLYR3 orthologs can be associated with loss of AM (Bouchiba et al 2022;Malkov et al 2016) and MtLYR8 (LYR-IB subclass) since, like LYR-IA, this subclass is absent in the non-AM species A. thaliana (Buendia et al 2018) and shown to be involved in AM in barley (Li et al 2022). Clearly, analysis of double, triple and higher order mutants are required to unravel possible LysM-RLK redundancy in AM.…”

“…We identified a high affinity (nM range) LCO binding protein in the PM fraction of cell cultures as the LYR-IIIA protein, MtLYR3 (Fliegmann et al 2013;Gressent et al 1999). MtLYR3 is located next to and in inverse orientation to MtNFP (Buendia et al 2018;Gough et al 2018), and legume orthologs of MtLYR3 bind LCOs (Bouchiba et al 2022;Malkov et al 2016). We also used a radiolabelled LCO-V ligand (LCO-V(C18:1,NMe, 35 S)), which has more similarity to a Myc-LCO than the S. meliloti NFs, to characterise LCO binding to the tomato and P. hybrida LYR-IA protein, and showed the relevance of these proteins for AM (Girardin et al 2019).…”

A receptor required for chitin perception facilitates arbuscular mycorrhizal associations and distinguishes root symbiosis from immunity

“…), and Parasponia andersonii ( P. andersonii ). 17 , 18 , 19 , 20 , 21 , 22 , 23 Besides CO, AMF also produce LCO, 8 and several LysM-RLKs have been shown to bind LCO in legumes 24 , 25 , 26 , 27 , 28 , 29 , 30 and act as pivotal components for LCO detection in the root nodule symbiosis (RNS), notably MtNFP / LjNFR5 and MtLYK3 / LjNFR1 in legumes M. truncatula and Lotus japonicus . 31 , 32 , 33 , 34 , 35 , 36 , 37 These LCO receptors, which are essential for the RNS, also contribute to the AMS, as evidenced by a quantitative reduction in AMF colonization in the cerk1 / nfp double mutant, as compared with cerk1 alone.…”

A new group of LysM-RLKs involved in symbiotic signal perception and arbuscular mycorrhiza establishment

“…Several LysM-RLK/Ps have been described as high affinity LCO or long-chain CO binding proteins [8][9][10][11][12][13][14][15][16][17] .…”