Molecular cloning and functional characterization of NtWRKY41a in the biosynthesis of phenylpropanoids in Nicotiana tabacum

Phenylpropanoids are important secondary metabolites that have multifaceted effects on plant growth, development, and environmental adaptation. WRKY41 has been shown to repress anthocyanins synthesis in Arabidopsis, but its full roles in regulating plant phenylpropanoids metabolism still remains to be further studied. Here, we cloned two NtWRKY41 genes from N. tabacum genome, and NtWRKY41a showed higher expression levels than NtWRKY41b genes in all the tobacco tissues examined. Overexpression and knock-out of NtWRKY41a gene revealed that NtWRKY41a promoted the biosynthesis of Chlorogenic acid (CGA) and lignin, but repressed the accumulation of scopoletin and flavonoids in tobacco. Transcriptome analysis found 7 phenylpropanoids related differentially expressed genes (DEGs) between WT and NtWRKY41a-OE plants, among which the transcription of NtCCoAOMT and NtHST was significantly induced by posttranslational activation of NtWRKY41a, while those of NtF6'H1 and NtGT3 was significantly repressed by NtWRKY41a. Chromatin immunoprecipitation and Dual-Luc assays further indicated that NtWRKY41a could bind to the promoter regions of these four genes to regulate their transcription. Moreover, ectopic expression of NtWRKY41a also promoted the transcription of several NtLOX and NtHPL genes, which encode key enzymes involved in the oxylipin pathway. Our findings revealed new functions of NtWRKY41a in modulating the distribution of metabolism flux in phenylpropanoids pathway, and provided a promising target for manipulating phenylpropanoids contents in tobacco.