Molecular Dissection Of Arabidopsis Endoplasmic Reticulum Protein SES1 In Regulating Salt Stress Tolerance

Soil salinization seriously affects the growth and development of plants,and becomes one of the major environmental factors in limiting crop yields.High salinity in soil causes ionic imbalance,osmotic stress and oxidative damage.Nearly 20%of the world's cultivated land and nearly half of irrigated area are affected by salt stress.Therefore,how to efficiently use the salinized soil,cultivate new salt-tolerant crops,become a global concern in the world.To survive,plants have evolved a number of different physiological and biochemical mechanisms to protect themselves from reducing the damage of salt stress.For example,the reconstruction of ions balance,biosynthesis of osmoprotectants,activation of antioxidant enzymes and regulation of genes expression,etc.At the same time,in recent years,studies have shown that endoplasmic reticulum also plays vital roles in signal transduction and integration under salt stress.Salt stress can disturb the normal protein folding process,cause unfolded protein accumulation in endoplasmic reticulum and lead to endoplasmic reticulum stress in plants.However,the reciprocally regulatory relationships between salt response and ER stress are still elusive.Through forward genetics,we isolated and identified SES1?SENSITIVE to SALT 1?in Arabidopsis,which plays important roles in salt stress resistance by alleviating salt induced ER stress.Our experimental results are as follows:?1?EMS mutagenesis mutant ses1-1 was hypersensitive to salt stress.By map-based cloning,we found the candidate gene termed SES1.SES1 can complement the salt sensitive phenotype of ses1-1 and the T-DNA knockout mutant ses1-2.?2?SES1 protein only exists in plants and contains a Saposin B domain.Mutated glutamic acid of ses1-1 is just located in this conserved region.There are altogether six proteins containing the Saposin B domain in Arabidopsis.Evolutionary relationships analysis showed that the remaining five proteins were far from SES1.The biological functions of SES1 orthologues are still not well characterized.?3?Overexpression of SES1 does not enhance the salt stress resistance of transgenic Arabidopsis.?4?In addition to NaCl,ses1-1 and ses1-2 also showed hypersensitivity to NaNO₃,KCl,KNO₃,mannitol,and LiCl,respectively,indicating that SES1 is required for both ionic and osmotic stress tolerance.?5?SES1 was ubiquitously expressed in root,shoot,rosette leaf,cauline leaf,flower,silique,and seed.Meanwhile,SES1 could be upregulated by salt stress.SES1 was located in the endoplasmic reticulum under normal or salt-stressed conditions.?6?The Na⁺content and expression of salt stress responsive genes in ses1 were not significantly different from that in wild type.RNA-seq analysis showed that differentially expressed genes involved in endoplasmic reticulum stress changed obviously in ses1 after salt treatment,indicating that ses1 was subjected to more severe endoplasmic reticulum stress after salt stress.Proteomic analysis showed that 14 ER proteins involved in protein processing and folding were significantly increased in ses1-2 after salt treatment,indicating more improperly folded proteins accumulation in ses1-2.Transmission electron microscopy showed significantly swollen endoplasmic reticulum lumen in ses1,and formed a lot of endoplasmic reticulum pools even showed tearing phenomena after salt stress treatment.?7?SES1 has the activity of molecular chaperone,which is involved in the folding of proteins.The fluorescence intensity of secGFP in ses1 was much higher than that in WT,and the fluorescence signal is mainly produced within the ER,suggesting the secretion of secretory proteins was severely blocked in ses1,which gave rise to ER stress.?8?SES1 was remarkably induced by exogenous tunicamycin treatment,and ses1 mutants showed sensitivity to TM.There is an ERSEL element within the promoter region of SES1.EMSA and Ch IP assay showed that ER stress sensor b ZIP17 can target ERSEL element to activates the expression of SES1.Moreover,the salt sensitivity of bzip17 could be rescued by enhancing the expression level of SES1.