Molecular Characterization Of Ethylene-responsive Transcription Factor AcERF1from Atriplex Canescens (Pursh) Nutt

Biotic and abiotic stress factors such as harmful microorganisms, salinity, heat, cold anddrought cause widespread crop losses throughout the world. It is estimated that every yearpathogen attacks causes$200billion crop losses throughout the world, and only20%ofagricultural land is immune to multiple stress (e.g., drought and salinity). The four-wing saltbush,Atriplex canescens, one of the most important halophyte, has been extensively recommended asan excellent phytoremediation plants in saline-alkali and heavy-metal contaminated land. In thisstudy, a new member (AcERF1) of ERF family was isolated from the A. canescens cDNA libraryand the molecular function and regulation mechanism were further analyzed. The experimentresults are following:1. The AcERF1contains a typical AP2domain and shares about20%homology with themembers of Group VII from Arabidopsis thaliana,2. Heterologous expression of the AcERF1gene in yeast significantly increased the ability oftransgenic yeast cells to adapt to and recover from exposure to to KCl, sorbitol and paraquat,AcERF1gene expression also provided osmotic, alkaline, and high temperature stress tolerancein yeast cells;3. Subcellular localization of an AcERF1/GFP fusion protein expressed in tobacco cellsshowed that AcERF1was localized in the nucleus. AcERF1was expressed in yeast to analyze itstranscriptional activation, and the results indicated AcERF1had transcriptional activation abilityin yeast which suggested it is a transcription activator;4. qRT-PCR result shows that drought, salinity, freezing and alkali could induce theexpression level of AcERF1gene in A. canescens. As an ethylene responsive factor, AcERF1wasextremely sensitive to the treatment of ethylene, moreover, AcERF1transcript level alsoaccumulated with application of exogenous SA, MeJA and ABA;5. Overexpression of AcERF1in A. thaliana showed increased resistance to Pseudomonassyringae pv tomato DC3000and Botrytis. cinerea. The expression level of pathogenesis-related gene in transgenic Arabidopsis was detected by qRT-PCR. The results indicated that the highexpression level of PR1, PR2, PR5, PR4, PR8, Pdf1.2and the interaction of AcERF1with ERF1,ERF3underlined the strong tolerance of transgenic line to PstDC3000and B. cinerea.6. AcERF1transgenic lines were hypersensitive to ABA during germination, seedlingestablishment and primary root elongation, and the stomatal movement also showed a sensitivebehavior; Furthermore, overexpression plants showed remarkably enhanced tolerance to droughtduring every stage. To reveal how AcERF1in transgenic plant enhances to drought-tolerance, weexamined the lower reactive oxygen species (ROS) levels (H2O2and O2.-) content,malondialdehyde (MDA) content, antioxidant enzyme activities(POD, SOD) and the expressionlevel of ROS-related and stress-responsive genes under osmotic stress. These results showed thatAcERF1overexpression in Arabidopsis enhances the transcript level of ROS-related and stress-responsive genes, which contribute to the higher relative water content (RWC) and superoxidedismutase (SOD) and catalase (CAT) activities in AcERF1transgenic line.