The Functions Of U-box Gene TaPUB1 In Plant Salt Tolerance Of Wheat

Wheat(Triticum aestivum L.)is a major food crop which is cultivated widely around the world.However,wheat often suffers from various stresses.The ubiquitin/26S proteasome system(UPS)is one of the most important protein turnover mechanisms in plants.It is mainly composed of Ub-activation enzyme(E1),Ub-conjugating enzyme(E2),Ub ligase(E3)and 26S proteasome and ubiquitin(Ub).UPS is involved in many processes,such as signal transduction,cell cycle and stress responses including abiotic stress response and plant defense.In our previous study,a wheat U-box E3 ubiquitin ligases gene TaPUB1 was obtained.To understand the function of TaPUB1 in wheat under salt stress,overexpressing and RNA interference transgenic wheat plants were generated.This study analyzed the salt resistance of transgenic wheat and its mechanism.The main results are as follows:(1)Analysis was performed by PCR and qRT-PCR,then we choose two overexpression and suppression lines which were used in the following experiments.(2)The salt tolerance of the different growth stages of transgenic and WT wheat was evaluated.The result show that the shoot and root growth of TaPUB1-OE were slightly inhibited compared with WT.However,the growth of TaPUB1-RNAi seeding were significantly inhibited.In addition,the TaPUB1-OE wheat kept higher photosynthesis ability compare to WT under salt stress.But the photosynthesis ability trends of TaPUB1-RNAi were opposite to TaPUB1-OE under salt stress.These results indicated that TaPUB1 played a role in the increasing salt tolerance in wheat(3)The accumulation and fluxes of Na~+and K~+was analyzed by ion toxicity.The results show that Na~+concentration in TaPUB1-OE were lower than WT under salt stress,while the K~+concentration in TaPUB1-OE were higher than WT.That indicated that the Na~+/K~+in TaPUB1-OE was lower than WT.But the Na~+/K~+in TaPUB1-RNAi was higher than WT under salt stress.In addition,we detected Na~+,K~+and H~+fluxes,and we found that TaPUB1 could regulate Na~+effluxes and K~+maintained which was corresponding to the result that TaPUB1 could regulate low Na~+and high K~+in wheat cells.(4)We analyzed the mechanism of salt tolerance of transgenic wheat by the physiology,biochemistry and molecular determinations.The result show that the TaPUB1-OE had higher activities of antioxidant enzyme compare with WT under salt stress,so the ROS content,MDA content and electrolyte leakage in TaPUB1-OE was low than WT.But the ROS content trends of TaPUB1-RNAi were opposite to TaPUB1-OE under salt stress.These results indicated that TaPUB1 regulates ROS accumulation via activities of antioxidant enzyme under salt stress,thereby contributing towards wheat salt tolerance.(5)Analyzed by detecting the expression of salt stress-associated,ion transport-associated and ROS scavenge-associated genes.It was found that the expression levels of most of the genes were consistent with previous phenotypic results.(6)Based on a Y2H screening assay,we selected TaMP as a candidate protein responsible for salt stress,the TaPUB1 protein interacting with TaMP in the nucleus.We generated transgenic Brachypodium distachyon plants overexpressing TaMP.The results suggest that TaMP-overexpressing Brachypodium distachyon are more sensitive to salt stress in root growth.In contrast to the expression patterns of TaPUB1,TaMP functions as a negative regulator of the salt stress response.In summary,our findings highlight a potential new route for fine-tuning the salt stress response in wheat via TaPUB1 and TaMP.TaPUB1 induces the expression and activity of ion transporters to regulate intracellular Na~+,K~+,and H~+fluxes,to maintain a lower cytosolic Na~+/K~+ratio under salt stress.Concurrently,TaPUB1 could also increase salt tolerance through increased expression of antioxidant genes and decrease the cellular levels of ROS under salt stress.Furthermore,TaMP acts as a negative regulator of the salt stress response,and interacts with TaPUB1 in the nucleus.