A method for functional testing constitutive and ligand-induced interactions of lysin motif receptor proteins
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(2020) 16:3 Li et al. Plant Methods https://doi.org/10.1186/s13007-020-0551-4
Open Access
METHODOLOGY
A method for functional testing constitutive and ligand‑induced interactions of lysin motif receptor proteins Chun‑Lian Li, De‑Xing Xue, Yi‑Han Wang, Zhi‑Ping Xie* and Christian Staehelin*
Abstract Background: Plant receptors with lysin motifs (LsyM) recognize microbial signals such as fungal chitin and lipochitooligosaccharidic Nod factors of nitrogen-fixing rhizobia. It is generally assumed that ligand-induced dimerization of LysM receptors is an essential step in activation of intracellular kinase domains and downstream signaling. Conse‑ quently, genes required for plant defense and establishment of symbiosis are expressed. We recently found that three LysM receptor proteins (namely LYK1, LYK4 and LYK5) of Arabidopsis thaliana form a tripartite receptor complex to perceive chitin. However, constitutive and ligand-induced interactions of LysM receptors generally remain difficult to be characterized. Results: Interactions between ectodomains of LYK1, LYK4 and LYK5 were investigated by a chimeric receptor approach using hairy roots of the legume Lotus japonicus. Synthetic receptor pairs consisting of a LYK ectodomain and the intracellular domain of a L. japonicus Nod factor receptor (NFR1 and NFR5, respectively) were tested for their capacity to activate expression of the symbiotic NIN (nodule inception) gene. The results indicated constitutive (LYK4ED–LYK4ED, LYK4ED–LYK5ED) and chitin-induced interactions (LYK1ED–LYK1ED, LYK1ED–LYK5ED) of the examined ectodomains. Conclusion: We present a method to functionally analyze constitutive and ligand-induced interactions of LysM-type proteins. Keywords: Chimeric receptor, Lotus japonicus, Lysin motif (LysM), LysM receptor-like kinase (LYK), Receptor function Background Plants possess specific receptors and signal cascades to alter gene expression in response to invading microbes. PAMPs (pathogen-associated molecular patterns) are conserved elicitor molecules such as fungal chitin and chitooligosaccharides, bacterial peptidoglycan, and flagellin [1–3]. PAMPs are typically perceived by corresponding PRRs (pattern recognition receptors) in plant plasma membranes. Many PRRs are receptor kinases *Correspondence: [email protected]; [email protected] State Key Laboratory of Biocontrol and Guangdong Key Laboratory of Plant Resources, School of Life Sciences, Sun Yat-Sen University, East Campus, Guangzhou 510006, Guangdong, China
consisting of an extracellular, ligand-binding ectodomain, a transmembrane domain, and an intracellular kinase domain. Some PRRs lack an intracellular kinase domain or possess an inactive kinase domain. This implies that these receptors require cooperation with another PRR or an adaptor kinase. PRR-mediated signaling results in activation of PAMP-triggered defense reactions (plant immunity) [1, 2, 4]. To recognize symbiotic microbes, host plants perceive specific microbial signals to activate expression of symbiosis-related plant
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