Functional Study Of MYB111 Regulates Salt Stress Response In Arabidopsis Thaliana

Abiotic stresses such as high salinity and drought seriously affect plant growth and crop yield.High salt stress caused ion totoxicity,osmotic stress and oxidative damage to plant cells.Therefore,the research on the important components involved in salt stress and functional mechanismsis is of great theoretical and practical significance to understand plant salt stress response mechanism,which is important for the rational utilization of genetic resources and the cultivation of crops with salt tolerance and high yield.MYB transcription factor is one of the largest transcription factor familys in plants,which plays important roles in plant response to biotic and abiotic stress.Studies have shown that MYB111 regulates the biosynthesis of flavonoids(secondary metabolite in phenylpropane pathway)in Arabidopsis thaliana.Flavonols play important roles in protecting plant oxidative damage under biotic and abiotic stresses.In this study,T-DNA insertion mutants of MYB111,Cas9-edited homozygotes and transgenic overexpression lines were used to the study of MYB111 function in plant response to abiotic stresses.A series of results were obtained.Through the identification of homozygous,vector construction and transformation screening,T-DNA insertion homozygous mutants(myb111),Cas9 editing lines of MYB111(myb111-d1)and transgenic overexpression lines(OE-2,OE-8)were obtained.Phenotypic identification showed that myb111 and myb111-d1 were higher sensitive to NaCl treatment during germination and seedling growth,and enhanced ROS production(hydrogen peroxide and superoxide anion)after subjected to salt treatment.By contrast,lines with MYB111 overxpression(OE-2,OE-8)were more tolerance to NaCl treatment than WT,and lower ROS accumulation recorded.myb111 and myb111-d1 demonstrated lower flavonol content,higher anthocyanin content,weaker ROS(O2-,·OH,DPPH)scavenging ability and lower antioxidant enzyme(SOD,POD,CAT)activity under salt stress,which were opposite in transgenic MYB111 overexpression seedlings compared with the corresponding control(WT).Those results indicate that MYB111 regulates NaCl-induced ROS accumulation under salt stress by regulating flavonoids biosynthesis and affecting autioxidant enzyme activity.qRT-PCR and tissue staining of the GUS(driven by the MYB111 promoter)transgenic lines revealed that the expression of MYB111 was induced by salt stress.The expression of key enzyme genes involved in flavonoid biosynthesis(CHS,CHI,F3H,FLS)and ROS scavenging enzyme(SOD,POD,CAT)are inhibited in myb111 under normal condition and salt stress.The deletion of MYB111 also produced a low expression level of salt stress response genes such as SOS1 during NaCl treatment,suggesting that MYB111 also regulates salt stress response through indirectly regulation of salt stress signal transduction pathways in Arabidopsis.Expression detection obtained from qRT-PCR and GUS staining indicated that the expression of MYB111 was in all detected tissues(including root,leaves,stem,flowers,siliques),especially in leaves.As an R2R3-MYB transcription factor,MYB111 is located in the nucleus and has transcriptional activation activity.Transcriptional activation roles of MYB111 to key enzyme genes involved in flavonols biosynthesis has been confirmed before,and the transcriptional activation regulation of MYB111 to other genes(such as SOD,CAT,POD,RCD1,etc)need to be further studied.In conclusion,our study showed that MYB111 regulates plant salt stress reponse and salt tolerance by impairing salt-induced ROS accumulation(regulating flavonol synthesis and influencing ROS scavenging enzyme gene expression and activity)and regulating salt stress signaling pathway.This study provides experimental investigation and theoretical basis for the analysis of the functional mechanism of MYB111 in response to abiotic stress.