Function Analysis Of Salix Linearistipularis SlRUB Gene In Response To Saline-alkali Stress

Salix linearistipularis is a perennial woody plant,which grows in soda and saline-alkali soils and has strong saline-alkali tolerance.Rubredoxin(RUB)is a class of iron-sulfur protein localized on the thylakoid membrane in chloroplasts.RUB is involved in SOR superoxide reduction,photosynthetic system assembly and repair,SOS signal transduction pathway,as well as ROS scavenging pathway,but the molecular mechanism of its response to saline-alkali stress is still unclear.In this study,We used physiological and molecular genetic strategies to analyze the salinealkali response function of the Sl RUB gene of Salix linearistipularis.The main findings include:The phylogenetic tree and amino acid sequence alignment analyses showed that RUB proteins from diverse plant species have four conserved cysteine(Cys)sites,which can form two pairs of disulfide bonds.If the conserved Cys in the Rubredoxin domain of the Sl RUB was changed to Ala by using molecular biology method,Sl RUB would lose the ability to form disulfide bonds and participate in redox process.However,the Cys site mutation did not affect the localization of Sl RUB in chloroplasts.The phenotype analysis of Sl RUB transgenic Arabidopsis thaliana seedlings showed that Sl RUB protein promotes rosette leaf growth and bolting.Compared with Cys203-Cys222,Cys200-Cys225 may play a more important role.Under saline-alkali stress conditions,the seed germination rate and chlorophyll content of Sl RUB-overexpressed Arabidopsis thaliana seedlings were increased.Bi FC analysis showed that Sl RUB interacted with At D1,suggesting that Sl RUB may enhance salt and oxidative tolerance in seedlings by regulating photosynthesis in Arabidopsis thaliana overexpressing Sl RUB.In addition,DAB and NBT staining assay showed that the mutant plants accumulated more ROS.The ROS accumulation in Sl RUB-overexpressed Arabidopsis thaliana was lower than that in the wild-type plants,and the ROS accumulation in the Sl RUB-complemented plants was also less than that in the mutant.Cys200-Cys225 may play a more important role in the ROS scavenging pathway.It is speculated that Sl RUB may improve plant salt tolerance by scavenging ROS.The above research results provide important information for in-depth understanding of the function of the halophyte Salix linearistipularis Sl RUB in the process of saline-alkali response.