Research On The Transcriptional Regulatory Mechanism Of BTH-induced Disease Resistance In Triticeae Crops

Systemic acquired resistance(SAR)is an inducible form of plant disease resistance that can generate broad-spectrum resistance to the secondary pathogens.In Arabidopsis thaliana,initial infection of pathogenic bacteria or treatment of BTH(benzothiadiazole)can induce the SAR.Meanwhile.SAR is associated with accumulation of plant hormone SA and transcriptional activation of PR gene.In A rabidopsis,NPR1 is a key transcriptional regulator of plant SAR.In response to the accumulation of plant hormone SA,NPR1 is translocated from cytoplasm to nuclei and interacts with a basic-region leucine zipper transcription factor(TGA2)to trigger the expression of PR(pathogenesis-related)genes expression.However,in Triticeae crops of wheat and barley,BTH-induced resistance(BIR)to various pathogens,including powdery mildew and leaf-rust,is not regulated by NPR1.In this study,RNA-seq analysis was performed on barley transgenic lines overexpressing wheat wNPR1 gene wNPR1-OE,knocking down barley HvNPR1 gene HvNPR1-Kd and wild-type plants,to characterize the role of NPR]in BIR and draft the expression profiles of WRKY transcription factor in a genome-wide level.The expression levels of NPR1,HvBCI1,HvBCI,HvBCI7,TaPR1a,TaPR2,and HvChit were further detected by qRT-PCR assay.Most of the BCI(barley chemical-induced)genes were induced in a NPR1-independent manner.Whereas,in barley transgenic lines wNPR1-OE,a few PR genes became more sensitive to BTH treatment.A large number of transcription factors,especially WRKY transcription factor,were significantly up-regulated upon BTH treatment,and the expression patterns of a few WRKY transcription factors were correlated with the expression of NPR1 transgene.Seven WRKY genes with significant up-regulation during BIR.including HvWRKY2,HvWRKY28,HvWRKY41,HvWRKY46,HvWRKY48,HvWRKY50,and HvWRKY52,were selected for further functional characterization.Transient expressions of these WRKY genes in wheat leaves using Agrobacterium indicated that HvWRKY41 and HvWRKY46 could significantly enhance the resistance of wheat to leaf rust virulent pathtype THTT.Meanwhile,all the candidate WRKY transcription factors were screened for their interactions with wNPR1 protein by yeast two-hybrid(Y2H)assay and none of them showed any direct association with wNPR1.Subcellular localization assay using GFP tag showed that both HvWRKY41 and HvWRKY46 were located in the nucleus.In conclusion,the presented study has provided a regulatory network of BIR and the identified key regulatory factors will be valuable genetic resources for the improvement of wheat broad-spectrum disease resistance.