Function Analysis of Wheat (Triticum aestivum) TaWRKY46 Gene in Mediating Salt Stress Tolerance in Transgenic Tobacco (Nicotiana tabacum)
JIANG Ming-Yue1,2, SU Xiao-Shuai1,2, ZHANG Bao-Hua1,2, LI Xiao-Juan1,2,*, XIAO Kai3,*
1 College of life Sciences, Hebei Agricultural University, Baoding 071001, China; 2 Key Laboratory of Plant Physiology and Molecular Pathology, Baoding 071001, China; 3 College of Agronomy, Hebei Agricultural University, Baoding 071001, China
Abstract：The members of the WRKY transcription factor (TF) family play important roles in mediating plant tolerance to salt stress. Previously, TaWRKY46, a member of WRKY gene family, was revealed to obviously respond to salt stress. Thus transgenic tobacco (Nicotiana tabacum) lines overexpressing this wheat gene were generated. The experiment indicated that TaWRKY46 targeted onto nucleus at subcellular level. Using the culture methods of vermiculite-based and Murashige & Skoog (MS) hydroponic solution, the phenotype of wild type (WT) and overexpression lines (OE) under salt stress treatment was investigated. OE1 and OE5, two OE lines overexpression TaWRKY46, displayed increased growth vigor and leaf area of plants, together with enhanced plant fresh weight and contents of soluble sugar and proteins upon salt stress with respect to WT. Assays on the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT), the enzymes worked as cellular protector, showed the higher activities in OE lines than those in WT plants. Reverse transcription PCR (RT-PCR) analysis indicated the expression levels of some protection enzymes mentioned above from roots were higher in OE1 and OE5 than those in WT under salt stress treatment. Further histochemical staining study via nitroblue tetrazolium (DAB) and diaminobenzidine (NBT) dyeing methods revealed that the accumulative amounts of O2-· and H2O2 were lower in the OE plants than that of WT plants. The expression analysis revealed that the pyrabactin resistance-like abscisic acid receptor gene (PYL8) and a suite of sucrose non-fermenting-1-related protein kinase 2 (SnRK2) family genes were upregulated in expression in the OE plants. Observation on stomata movement showed that the OE plants possessed enlarged stomata aperture and increased closure rate upon salt stress compared with the WT plants. Using yeast-two hybridization assay, TaWRKY46 was identified to be interacted with TaSAP1-1, a stress-related protein in T. aestivum. Therefore, TaWRKY46 played an important role in salt tolerance through enhancing osmotic regulation and protective enzyme system, crossing with ABA signaling pathway, as well as initiating protein interaction. The results enrich the knowledge as to wheat plants coping with salt stress and provide theoretical guidance for breeding stress tolerant cultivars of crops.
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