Cloning And Functional Analysis Of The OsSTG1 Gene For Salt Tolerance At The Germination Stage Of Rice (Oryza Sativa L.)

Soil salinity,as one of the most brutal abiotic stresses,negatively influences the survival,biomass production and yield of crop.Rice(Oryza sativa L.)is sensitive to salt stress,especially at the germination and seedling stages.High salt stress results in the reduction of germination and seedling survival rate.Isolating salt tolerant genes and researching on the mechanism of salt tolerance play the most important role in developing salt tolerant cultivars of rice.Several years ago,a set of introgression lines,based on an accession of Yuanjiang common wild rice(YJCWR,Oryza rufipogon Griff.)in the back ground of indica cultivar teqing,were used to evaluate for the salt sensitivity at the germination stage and one introgression line YIL3 was identified.Then a major QTL(qRGE5)for salt tolerance at the germination stage was mapped on chromosome 4.In this study,a segregation population derived from the cross between YIL3 and Teqing was used to delimit qRGE5 to a 9 kb region.Within this region,there is only one candidate gene LOC_Os04g53190.To verify whether LOC_Os04g53190 controls the salt tolerance at the germination stage of rice,an over-expression(OX)vector was generated and introduced into YIL3.Six independent OX-transgenic lines displayed the salt tolerant phenotype like Teqing with the up-regulated expression of LOC_Os04g53190.Meanwhile,an RNAi construct was introduced into Teqing.Five independent RNAi-transgenic lines exhibited the salt sensitive phenotype like YIL3 with the down-regulated expression of LOC_Os04g53190.These results showed that LOC_Os04g53190 could control the salt tolerance at the germination stage of rice,named OsSTG1(Salt Tolerance at the Germination Stage 1).Analysis of the OsSTG1 expression patten by RT-PCR showed that the expression of OsSTG1 was up-regulated under the salt stress.The results of the RT-PCR and GUS staning indicated that OsSTG1 was expressed in various tissues at the different stages of rice,especially highly expressed in embryo tips at the germination stage.Transient expression of the construct OsSTG1-GFP showed that STG1-GFP localized to the cytoplasm in the epidermal cells of onions.Analysis of the OsSTG1-encoded protein function revealed that the OsSTG1 gene might encode a polyamine oxidase(PAO)in the cytoplasm,involved in polyamines(PA)catabolism,which could oxidate spermine(Spm)to spermidine(Spd),leading to the increase of Spd and PA contents.Meanwhile,PAO oxidated Spm with the production of hydrogen peroxide(H2O2),which was related to the adaption of plants to salt stress.To explore how OsSTG1 increases the salt tolerance at the germination stage,several analyses were performed.The results showed that the elevated expression of OsSTGl enhanced not only the expression of several defense genes in response to salt stress,but also the expression of genes enconding reactive oxygen species(ROS)scavenging enzymes,leading to increasing activities of these enzymes and reducing the H2O2 accumulation in the embryos to maintain ROS homeostasis,which consequently increased the Na+efflux to reduce Na+content in embryos and maintained ion homeostasis for healthy shape of embryo tips,that resulted in the better germination of seeds and elongation of coleoptiles at the germination stage.