Response Of Grape WRKY30to Biotic And Abiotic Stresses

WRKY30transcription factor (TF) was clonned from downy mildew (DM) immuned grape cultivar Muscadine rotundifolia ’Noble’, highly DM resistant grape cultivar Vitis amwensis ’Zuoshan1’, and susceptible grape cultivar Vitis vinifera’Thompson Seedless’, respectively. These WRKY30TF contain1050bp basic group, coding349amino acids. All of them have nuclear localization sequence ’KKRK’and the III type zinc finger ’CCHC’ Similarity of among these WRKY structural domain is100%. Sequnce alignment and phylogenetic analysis show that grape WRKY30has low similarity and far genonomy with those studied grape WRKY transcription factors. Subcellular localization of WRKY30reveals that the WRKY30is located in the nucleus, and it possibly works as DNA-binding TF.Grape WRKY30was transformed to Arabidopsis to study its function. The WRKY30had no effect on drought tolerance and driage of leaves, but increased the sensibility of Arabidopsis to high salt concentration by suppressing the expression of ascorbate peroxidase (AtAPX), catalase (AtCAT) and glutathione-S-transferase (AtGST). After infection of Peronospora parasitica, the expression levels of pathogenesis-related genes (AtPR1, AtPR4, AtPR5, AtPDF1.2) and phytohormone related genes increased in transgenic Arabidopsis. The result indicated that WRKY30induced the expression of AtPRS by salicylic acid (SA) signaling pathway, and induced AtPDF1.2by jasmonic acid (JA) rather than ethylene (ET) signaling pathway. Grape WRKY30TFs were induced by downy mildew pathogen, of which the highest induction was observed in V. Amurensis’Zuoshan1’. The base line expression levels of WRKY30in disease-resistant grape varieties were much higher than those in the susceptible varieties. After abscisic acid (ABA), ethephon, methyl jasmonate (MeJA) and SA treatment, WRKY30TFs were either induced or suppressed among different grape varieties.To conform the PR genes regulated by WRKY30, prokaryotic expression vector pET30a_WRKY30was constructed to produce WRKY30protein. After purification, WRKY30were detected by electrophoretic mobility shift assays, showing that WRKY30could regulate the promoter with W-box. The candidate genes thaumatin-like proteins (TLP2-7, TLP2-8, TLP8-1, TLP8-3, TLP13-1and TLP18) were selected by promoter analysis. TLP2-7and TLP2-8were upregulated in ’Zuoshan1’after Plasmopara viticola infection. After instantaneous conversion of pH7WG2D_WRKY30to ’Thompson Seedless’ leaves, TLP2-7and TLP2-8were induced. After inoculation with conidiospore, the growth of P. viticola was suppressed and the expression level of TLP2-7increased. Therefore, WRKY30may increase the resistance of transgenic leaves by upregulating the expression of TLP2-7.