Study On The Functions Of Glutamate Receptor OsGLR3.4 In Plant Growth And Mechanical Wounding Stress Response In Rice

Rice is the main food crop in my country,and plant height and plant architecture are important agronomic traits that determine crop yield.In mammals,ionotropic glutamate receptors（i Glu Rs）are involved in the transmission of neural signals and the induction of neuron development.Glutamate receptor-like channels（GLRs）,the homologues of i Glu Rs in plants,have been shown to play important roles in various Ca²⁺-mediated plant-specific developmental processes and physiological responses.However,the functions of GLRs in plant height and plant architecture development and in response to environmental stress are still lacking systematic studies.Therefore,in this study,the functions of OsGLR3.4 in the development of plant height and plant architecture and the response to wounding were studied in rice,the monocot model plant.The main research work is summarized as follows:1.OsGLR3.4 plays crucial roles in brassinosteroid（BR）-mediated plant growth and development in rice.The loss-of-function mutants osglr3.4 exhibit the phenotypes such as dwarfism and reduced leaf inclination,accompanied by reduced cell length in the internode and lamina joint.osglr3.4 mutants have the reduced sensitivity to exogenous brassinosteroid（2,4-epibrassinolide,eBL）,indicating that they have typical phenotypes similar to BR signaling-defective mutants.These results demonstrate that OsGLR3.4 is involved in BR-mediated plant growth and development in rice.2.OsBZR1 directly regulates the transcription of OsGLR3.4.Electrophoretic mobility shift assays（EMSA）and transient transactivation assays indicated that OsGLR3.4 is a downstream target gene of OsBZR1,a key transcription factor in the BR signaling pathway,demonstrating that OsBZR1 activates the transcription of OsGLR3.4.3.A subset of amino acids can activate the ion channel activity of OsGLR3.4.Transmembrane Ca²⁺influx in wild-type coleoptile epidermal cells activated by a subset of amino acids is significantly reduced in osglr3.4 mutants,indicating that OsGLR3.4mutation reduces the cellular responses to multiple amino acids as agonists.These amino acids can activate the Ca²⁺channel activity of OsGLR3.4,thereby opening the Ca²⁺channel and inducing a transient influx of extracellular Ca²⁺,resulting in the cytoplasmic Ca²⁺concentration([Ca²⁺]_cyt)to increase instantaneously.Therefore,the OsGLR3.4 channel protein has a broad agonist spectrum.4.OsGLR3.4 is essential in Ca²⁺-mediated cytoskeleton microfilament arrangement and vesicle trafficking.The study of rice internode cells showed that the internode cell length of osglr3.4 mutants was reduced,accompanied by the disordered cytoskeleton microfilament arrangement and the blocked vesicle efflux.Application of La Cl₃,the Ca²⁺channel blocker,to wild-type internode cells caused the similar physiological response to the osglr3.4 mutant cells,indicating that OsGLR3.4-mediated[Ca²⁺]_cyt enhancement is indispensable for the correct arrangement of cytoskeleton microfilament and vesicle transport,thus maintaining cell elongation.5.OsGLR3.4 plays an important role in Ca²⁺-mediated systemic wounding signaling transmission from the mechanical injured root to the uninjured shoot.After root injury,the signal intensity of slow wave potentials（SWPs）induced by root injury and transmitted to leaves was significantly weakened,and the expression of JA-responsive gene also decreased in osglr3.4 mutants.Meanwhile,the sensitivity of osglr3.4 mutants to exogenous JA（methyl jasmonate,Me JA）was reduced,and OsGLR3.4 expression was also significantly reduced in the JA synthesis-deficient mutant oslox1,indicating that OsGLR3.4 is required for Ca²⁺-mediated systemic wounding signaling transmission.6.BRs synergistically regulate OsGLR3.4-mediated systemic defense responses in rice.eBL treatment increased OsGLR3.4 expression in wild type but not in the JA synthesis-deficient mutant oslox1.Meanwhile,eBL treatment increased the intensity of SWPs and the expression of JA-responsive gene in leaves in root-injured wild type,but not in root-injured osglr3.4 mutants,suggesting that BRs synergistically regulate OsGLR3.4-mediated systemic defensive response in rice.Collectively,our study elucidates the mechanism by which OsGLR3.4 mediates cell elongation and long-distance systemic injury signaling transmission in rice,and provides new insights into the roles of GLRs in plant growth and development and systemic defense responses.