Powdery mildew (PM), caused by the fungal pathogen Erysiphe necator, is one of the most destructive diseases of grapevine (Vitis vinifera and other Vitis spp). Resistance to PM is an important goal for cultivar improvement, and understanding the underlying molecular mechanisms conditioning resistance is critical. Here, we report that transgenic expression of the WRKY transcription factor gene VqWRKY31 from the PM-resistant species Vitis quinquangularis conferred resistance to powdery mildew in V. vinifera through promoting salicylic acid signaling and specific metabolite synthesis. VqWRKY31 belongs to the WRKY IIb subfamily, and expression of the VqWRKY31 gene was induced in response to E. necator inoculation. Transgenic V. vinifera plants expressing VqWRKY31 were substantially less susceptible to E. necator infection, and this was associated with increased levels of salicylic acid and reactive oxygen species. Correlation analysis of transcriptomic and metabolomic data revealed that VqWRKY31 promoted expression of genes in metabolic pathways and the accumulation of many disease resistance-related metabolites, including stilbenes, flavonoids, and proanthocyanidins. In addition, results indicated that VqWRKY31 can directly bind to the promoters of two structural genes in stilbene synthesis, STS9 and STS48, and activate their expression. Based on our results, we propose a model where VqWRKY31 enhances grapevine PM resistance through activation of salicylic acid defense signaling and promotion of specific disease resistance-related metabolite synthesis. These findings can be directly exploited for molecular breeding strategies to produce PM-resistant grapevine germplasm.

Anthocyanins are plant secondary metabolites that have a variety of biological functions, including pigmentation. The accumulation of anthocyanins is regulated by both transcriptional activators and repressors. Studies have shown that the bZIP family act primarily as positive regulators of anthocyanin biosynthesis, but there are few reports of negative regulation. Here, we report that a grapevine (Vitis vinifera) bZIP gene from group K, VvbZIP36, acts as a negative regulator of anthocyanin biosynthesis. Knocking-out one allele of VvbZIP36 in grapevine utilizing the CRISPR/Cas9 technology promoted anthocyanin accumulation. Correlation analysis of transcriptome and metabolome data showed that, compared with wild type, a range of anthocyanin biosynthesis genes were activated in VvbZIP36 mutant plants, resulting in the accumulation of related metabolites, including naringenin chalcone, naringenin, dihydroflavonols and cyanidin-3-O-glucoside. Furthermore, the synthesis of stilbenes (α-viniferin), lignans and some flavonols (including quercetin-3-O-rhamnoside, kaempferol-3-O-rhamnoside and kaempferol-7-O-rhamnoside) was significantly inhibited and several genes linked to these metabolism, were down-regulated in the mutant plants. In summary, our results demonstrate that VvbZIP36, as a negative regulator of anthocyanin biosynthesis, plays a role in balancing the synthesis of stilbenes (α-viniferin), lignans, flavonols and anthocyanins.

Summary Powdery mildew (PM) is a severe fungal disease of cultivated grapevine world‐wide. Proanthocyanidins (PAs) play an important role in resistance to fungal pathogens; however, little is known about PA‐mediated PM resistance in grapevine. We identified a WRKY transcription factor, VqWRKY56, from Vitis quinquangularis, the expression of which was significantly induced by PM. Overexpression (OE) of VqWRKY56 in Vitis vinifera increased PA content and reduced susceptibility to PM. Furthermore, the transgenic plants showed more cell death and increased accumulation of salicylic acid and reactive oxygen species. Transient silencing of VqWRKY56 in V. quinquangularis and V. vinifera reduced PA accumulation and increased the susceptibility to PM. VqWRKY56 interacted with VqbZIPC22 in vitro and in planta. The protein VqWRKY56 can bind to VvCHS3, VvLAR1, and VvANR promoters, and VqbZIPC22 can bind to VvANR promoter. Co‐expression of VqWRKY56 and VqbZIPC22 significantly increased the transcript level of VvCHS3, VvLAR1, and VvANR genes. Finally, transient OE of VqbZIPC22 in V. vinifera promoted PA accumulation and improved resistance to PM, while transient silencing in V. quinquangularis had the opposite effect. Our study provides new insights into the mechanism of PA regulation by VqWRKY56 in grapevine and provides a basis for further metabolic engineering of PA biosynthesis to improve PM resistance.

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