Mechanisms Of Pattern-Recognition Receptors Regulating Plant Immunity

Plants have evolved multiple layers of immunity to fend off pathogen attacks.Cell-surface localized pattern-recognition receptors(PRRs)sense conserved pathogen-associated molecular patterns(PAMPs)to mediate the pattern-trigged immunity(PTI).The best-studied PRRs include FLS2 and EFR,two Arabidopsis leucine rich repeat receptor kinases(LRR-RLKs)that recognize a conserved N-terminal 22 amino acid sequence of the bacterial flagellin and an acetylated N-terminal 18 amino acid sequence of bacterial elongation factor TU,respectively.The lysine motif(LysM)-containing RLK AtCERKl from Arabidopsis and its rice ortholog OsCERKl can bind chitin,which is a polymeric N-acetyl-D-glucosamine(NAG)and a major component of fungal cell wall.In rice,chitin perception also requires CEBiP,a LysM-containing receptor-like protein(RLP),which likely mediates immune signaling by an.association with OsCERK1.In addition,the Arabidopsis LysM RLPs,LYM1 and LYM3,and their orthologs in rice,LYP4 and LYP6,are responsible for the detection of peptidyl-glycan(PGN),a major cell wall component of gram negative bacteria.Furthermore,PEPR1 and PEPR2,two closely-related LRR RLKs from Arabidopsis,perceive a class of endogenous peptides called Peps,which are often produced during pathogen infection or wounding and are thus dubbed damage-associated molecular pattern(DAMP).In this study,we focus on mechanisms by which AtCERKl and PEPR1/PEPR2 mediate plant immunity.AtCERK1 contains an ectodomain(ECD)with three LysMs,a transmembrane domain(TM)and an intercellular serine/threonine kinase domain(KD).AtCERK1 senses chitin through the LysMs,but the underlying mechanism has remained unknown.Here,we demonstrate that chitin induces AtCERKl dimerization,and this is essential for the activation of plant immunity.Structural data of an AtCERK1-ECD-chitin pentamer complex suggest that higher oligomeric chitins such as(NAG)8 can simultaneously bind two or more AtCERKl molecules and that the clustered AtCERKl may be important for signaling.Indeed,crab shell chitin and(NAG)8 can induce the dimerization of AtCERKl-ECD and full-length AtCERKl both in vitro and in vivo.A138H,a mutation at the chitin-binding site of AtCERKl,attenuated the chitin-and(NAG)8-induced AtCERKl dimerization and FRK1 induction.Importantly,the chitin octamer-induced AtCERKl-ECD dimerization was attenuated by the addition of chitin pentamer in a dose-dependent manner.This is accompanied by a reduction of plant immune responses,such as FRK1 expression and reactive oxygen species.Furthermore,overexpression of AtCERKl-ECD prevented the formation of a productive AtCERKl dimer and abolished the chitin-induced AtCERKl phosphorylation and FRK1 expression.Together these data demonstrate that chitin-induced AtCERKl dimerization is critical for its activation.In plants,the defense phytohormones ethylene,jasmonate,and salicylate and DAMPs are induced to amplify immune responses upon the perception of pathogens.BIK1,a cytoplasmic receptor-like kinase,is a key component of PTI signaling.Surprisingly,the bikl Arabidopsis plant has been shown to display reduced sensitivity to ET.Here,we show through genetic and biochemical analyses that PEPRs and BIK1 mediate the ethylene-induced plant immunity.BIK1 and its close homolog PBL1 specially interact with the PEPR1 kinase domain to mediate the Pep-signal transduction.In vitro data showed that PEPR1,and likely PEPR2,directly phosphorylates BIK1.Like the bikl mutant,the pepr1/pepr2 double mutant etiolated seedlings displayed reduced sensitivity to ACC,as indicated by elongated hypocotyls.Accordingly,the ACC-induced plant resistance to Botrytis cinerea and the defense related genes expression were compromised in peprl/pepr2 and bikl mutants,reenforcing an important role of PEPRs and BIK1 in ET-mediated defense signaling.Pep treatment partially mimicked the ET-induced inhibition of hypocotyl elongation and root growth in a PEPRs-and BIK1-dependent manner.Furthermore,ACC and Pepl induced BIK1 phosphorylation in a PEPR-dependent manner,indicating that PEPRs are responsible for BIK1 phosphorylation in vivo.The Pepl-induced BIK1 phosphorylation,seedling growth inhibition,and defense-related genes expression occurred independent of EIN2 and EIN3/EIL1,which are key components of the canonical ethylene pathway,a finding consistent with the notion that Peps act downstream of EIN2,EIN3,and EIL1.Together our data shed light on a new mechanism by which ET and DAMP amplify plant immune responses.Together these data uncover two new mechanisms of plant immunity,namely the triggering of plant immunity by the chitin-induced dimerization of AtCERKl and the ethylene-induced immunity mediated by PEPRs and BIKl.