Structural and biochemical basis of FLS2-mediated signal activation and transduction in rice

The receptor-like kinase FLAGELLIN-SENSITIVE 2(FLS2)functions as a bacterialflagellin receptor local-ized on the cell membrane of plants.In Arabidopsis,the co-receptor BRI1-ASSOCIATED RECEPTOR KI-NASE 1(BAK1)cooperates with FLS2 to detect theflagellin epitopeflg22,resulting in formation of a signaling complex that triggers plant defense responses.However,the co-receptor responsible for recog-nizing and signaling theflg22 epitope in rice remains to be determined,and the precise structural mecha-nism underlying FLS2-mediated signal activation and transduction has not been claried.This study pre-sents the structural characterization of a kinase-dead mutant of the intracellular kinase domain of OsFLS2(OsFLS2-KDD1013A)in complex with ATP or ADP,resolved at resolutions of 1.98 A˚and 2.09 A˚,respectively.Structural analysis revealed that OsFLS2 can adopt an active conformation in the absence of phosphorylation,although it exhibits only weak basal catalytic activity for autophosphorylation.Subse-quent investigations demonstrated that OsSERK2 effectively phosphorylates OsFLS2,which reciprocally phosphorylates OsSERK2,leading to complete activation of OsSERK2 and rapid phosphorylation of the downstream substrate receptor-like cytoplasmic kinases OsRLCK176 and OsRLCK185.Through mass spectrometry experiments,we successfully identied critical autophosphorylation sites on OsSERK2,as well as sites transphosphorylated by OsFLS2.Furthermore,we demonstrated the interaction between OsSERK2 and OsFLS2,which is enhanced in the presence offlg22.Genetic evidence suggests that OsRLCK176 and OsRLCK185 may function downstream of the OsFLS2-mediated signaling pathway.Our study reveals the molecular mechanism by which OsFLS2 mediates signal transduction pathways in rice and provides a valuable example for understanding RLK-mediated signaling pathways in plants.

Structural and biochemical basis of Arabidopsis FERONIA receptor kinase-mediated early signaling initiation

Accumulating evidence indicates that early and essential events for receptor-like kinase(RLK)function involve both autophosphorylation and substrate phosphorylation.However,the structural and biochemical basis for these events is largely unclear.Here,we used RLK FERONIA(FER)as a model and crystallized its core kinase domain(FER-KD)and two FER-KD mutants(K565R,S525A)in complexes with ATP/ADP and Mg^(2+) in the unphosphorylated state.Unphosphorylated FER-KD was found to adopt an unexpected active conformation in its crystal structure.Moreover,unphosphorylated FER-KD mutants with reduced(S525A)or no catalytic activity(K565R)also adopt similar active conformations.Biochemical studies revealed that FER-KD is a dual-specificity kinase,and its autophosphorylation is accomplished via an intermolecular mechanism.Further investigations confirmed that initiating substrate phosphorylation requires autophosphorylation of the activation segment on T696,S701,and Y704.This study reveals the structural and biochemical basis for the activation and regulatory mechanism of FER,providing a paradigm for the early steps in RLK signaling initiation.