The Functional Identification Of Transcription Factor ERF72 In Arabidopsis And Wheat In Resistance To Waterlogging Stress

Waterlogging stress seriously affects the growth and development of plants,and even leads to plant death.Therefore,it is of great significance to study the mechanisms of plant resistance to waterlogging stress for crop genetic improvement.The transcription factor ERF-VII of Arabidopsis is an ethylene-responsive factor that plays an important role in plant resistance to waterlogging stress.In this study,Arabidopsis wild-type(Col),mutants atg10,erf72,erf74 and erf75 were used as experimental materials.Different time(0 h,4 h,8 h,12 h and 24 h)were set up for waterlogging(WL)treatment,and normal growth Arabidopsis was used as the control(CK),to study the changes of the expressi on levels of hypoxia responsive genes(HRGs),ethylene responsive factor ERF-VII and autophagy related genes(ATGs)in Arabidopsis,the interaction between ERF72 and ATG10 p,ERF74p,ERF75 p,and the effects of waterlogging stress on the growth state and physiological indicators of Arabidopsis.In order to explore the mechanism of ERF72 regulating the response to waterlogging stress.Then,9 wheat Ta ERF-VII were aligned by BALSTP,and their evolutionary relationships,protein conserved domains,tissue expression patterns and expression patterns under waterlogging stress were analyzed.The main results are as follows:1)Waterlogging stress can significantly induce the expression of hypoxia responsive genes HRGs in Col.and then respond to waterlogging stress.2)Waterlogging stress induced the up-regulation of ERF-VII subfamily genes in Arabidopsis,among which ERF72 was up-regulated most significantly.Under waterlogging stress,seed germination of Arabidopsis was inhibited,and the germination rate of erf72 seeds was significantly lower than that of Col.Waterlogging stress also resulted in shortened root length,stunted leaf development and decreased chlorophyll content,among which the growth inhibition of erf72 was aggravated.3)Under waterlogging stress,the genes expression levels of ERF74 and ERF75 in mutant erf72 were significantly decreased.Yeast one-hybrid(Y1H)and GUS staining experiments showed that the transcription factor ERF72 could bind to the promoter regions of ERF74 and ERF75.ROS staining showed that the content of reactive oxygen species in erf72 was significantly higher than that of wild type.It indicated that ERF72 may participate in the regulation of ROS by regulating the transcription of ERF74 and ERF75.4)The expression levels of autophagy related genes ATGs(ATG2,ATG7,ATG8 e and ATG10)increased under waterlogging stress,however,the gene expression level of ATG10 in mutant erf72 was always lower than that in wild type.Subcellular localization showed that both ERF72 and ATG10 were localized in the nucleus.Y1 H and GUS staining experiments revealed that ERF72 was involved in the transcriptional regulation of autophagy related gene ATG10.It indicates that ERF72 may regulate the occurrence of autophagy by regulating the transcription of the autophagy gene ATG10.5)9 Ta ERF-VII proteins with high amino acid sequence similarity to At ERF-VII were aligned from the wheat genome.Evolutionary analysis with At ERF-VII revealed that Ta RAP1 A,Ta RAP1 B and Ta RAP1 D are most closely related to At ERF72.Analysis of protein conserved domains shows that all 9 Ta ERF-VII proteins have a highly conserved AP2 domain located in the middle of the protein.The tissue expression pattern analysis showed that 9 Ta ERF-VII genes showed strong tissue specificity a nd were mainly highly expressed in roots and seeds.The results of expression level analysis under waterlogging stress showed that,except for Ta RAP5 D and Ta HRE7 D,which did not change significantly under waterlogging stress,the expression levels of other Ta ERF-VII genes increased significantly and reached their peaks at 4 h of waterlogging stress.The above results demonstrate that the wheat Ta ERF-VII genes,like the At ERF-VII genes,can respond to waterlogging stress.In summary,waterlogging stress can seriously affect the germination and growth of Arabidopsis.ERF72 may respond to waterlogging stress by participating in the generation of ROS and autophagy.And the mutation of ERF72 leads to reduced resistance to waterlogging stress in Arabidopsis.It was demonstrated that the transcription factor ERF72 plays an important role in Arabidopsis resistance to waterlogging stress.Preliminary study results of 9 ERF-VII subfamily genes in wheat indicated that Ta ERFVII genes responded to waterlogging stress,which provided a theoretical basis for the genetic improvement of wheat.