The HSFA2-HSFB2a-WRKY10 transcriptional cascade mediates thermotolerance across grape germplasms

Grapevines (Vitis vinifera) are economically important crops increasingly threatened by climate change-induced heat stress. Wild species such as V. davidii display markedly higher thermotolerance than the widely cultivated V. vinifera. Our previous research indicated that heat shock transcription factor A2 (VdHSFA2) from V. davidii confers higher thermotolerance than VvHSFA2 from V. vinifera, with HSFB2a as a potential downstream target of HSFA2. However, the precise mechanism by which HSFA2 modulates thermotolerance is not fully understood. Here, we demonstrated that HSFA2 directly activates HSFB2a, which in turn represses the transcription factor WRKY10. DAP-seq, yeast one-hybrid, and electrophoretic mobility shift assays confirmed HSFB2a binds to the WRKY10 promoter, and functional assays indicated that WRKY10 diminishes thermotolerance by repressing L-ascorbate peroxidase 3 (APX3) and 18.1 kDa class I heat shock protein (HSP18.1). However, HSFA2 does not bind to the WRKY10 promoter, and HSFA2, HSFB2a, and WRKY10 do not physically interact with one another. Transient expression in grapevine plants and stable transformation of suspension cells demonstrated that HSFB2a enhances, whereas WRKY10 reduces, thermotolerance-associated physiological performance. Notably, the V. davidii HSFB2a promoter is more active than that of V. vinifera HSFB2a, potentially contributing to species-level differences in thermotolerance. Our findings reveal the HSFA2-HSFB2a-WRKY10 transcriptional cascade modulating heat stress responses in grapevine, providing molecular targets for breeding heat-resilient cultivars.