Functional Analyses Of Receptor-like Kinase GSO1 In Regulating Salt Stress Response In Arabidopsis

High soil salinity is one of the major abiotic stresses affecting the global agriculture.Generally,plant salt stress can be subdivided into early-occurring osmotic stress and accumulating ionic stress.Increased soil salt concentrations(especially sodium)decrease the ability of plant to take up water,and excess sodium ions(Na+)taken by roots influence plant growth by impairing metabolic processes and decreasing photosynthetic efficiency.To respond to salt stress,plants have evolved numerous critical processes to limit toxic salt uptake and minimize the harmful effects of salt stress by exclusion of salt from cells and by compartmentalization of salt into vacuoles.To date,how the salt stress is sensed by plants,and whether plants have a Na+sensor or receptor remain elusive.Receptor-like kinases(RLKs)are better candidates as sensors to perceive environmental signals.To explore whether the RLKs involve in salt stress response,we screened a collection of T-DNA insertion mutants of Arabidopsis receptor-like kinase and isolated a salt-sensitive mutant,gsol.The GASSHO1(GSO1)gene encodes a typical LRR-RLK.Previous studies have reported that both GSO1 and GSO2 function in the regulation of embryos development.In this study,we found the salt-sensitive phenotype of gsol mutant in a GSO2 independent manner.The tissue specificity analysis of GSO1 expression showed that GSO1 was expressed mainly in the root.Under salt stress,the gsol mutant accumulated more Na+in root and shoot compared to the wild type,which indicates that GSO1 involves in Na+transport between root and shoot.To further study the molecular mechanism of GSO1 involved in plant salt response,we identified a putative GSO1 interacting protein,SOS2,by mass spectrum,and confirmed the interaction by Co-IP and BiFC.Using the reconstituted SOS system in yeast,we found that GSO1 activated the Na+transport activity of SOS1 via SOS2,but this process was independent of Ca2+ binding protein SOS3.The dead-kinasc of GSO1 could not activate the SOS pathway in yeast,which indicates that the kinase activity of GSO1 is essential for the activation of SOS pathway.GSO1 and SOS2 both are active kinases in vitro which have auto-and trans-phosphorylation activity.Our in vitro kinase assay showed that GSO1 phosphorylated SOS2,but could not be phosphorylated by SOS2.Compared to wild type,the kinase activity of SOS2 in the gsol mutant decreased when exposed to the NaCl treatment.These results suggest that GSO1 regulates the kinase activity of SOS2 both in vitro and in vivo.By mass spectrometer analysis,we found that GSO1 phosphorylated the Thr16 of SOS2.The mutations in Thr16 altered the SOS2 kinase activity.Genetic analysis of the gsol sos2,gsol sos3 and gsol scahp8 double mutants showed that GSO1 genetically interacted with SOS2,and was independent of SOS3 and SCaBP8 in modulating SOS pathway activity.Interestingly,expression of several salt stress-responsive genes decreased in the gsol mutant,which suggested that GSO1 may play an important role in the upstream of salt stress response.In summary,the receptor-like kinase GSO1 is a positive regulator in the Arabidopsis salt stress response.Our results demonstrate that GSO1 interacts with and activates the protein kinase SOS2 to regulate the Na+ transport of SOS1.GSO1 also regulates the expression of a series of downstream stress-responsive genes.This study uncovers a new Ca2+ independent pathway in regulating SOS pathway and provides a clue for a better understanding of sodium transport in plants.