Glutathione transferase VvGSTU60 is essential for proanthocyanidin accumulation and cooperates synergistically with MATE in grapes

Proanthocyanidin, synthesized in the endoplasmic reticulum and stored in vacuoles, is key to grape and wine quality. Glutathione S-transferase (GST) plays a crucial role in proanthocyanidin accumulation. However, little is known about the mechanisms of GSTs in the process. Here, we found that a TAU-type GST VvGSTU60 is required for proanthocyanidin accumulation in Vitis vinifera. Gene expression analysis revealed a favorable correlation between the expression pattern of VvGSTU60 and proanthocyanidin accumulation in the seed of V. vinifera. We discovered that the overexpression of VvGSTU60 in grapes resulted in a significant increase in proanthocyanidin content, whereas the opposite effect occurred when VvGSTU60 was interfered with. Biochemical analysis indicates that VvGSTU60 forms homodimers and heterodimers with VvGST1. Interestingly, we also found that VvGSTU60 interacts with VvDTX41B, a MATE transporter protein localized on the tonoplast. Heterologous expression of VvDTX41B in the Arabidopsis tt12 mutant rescues the proanthocyanidin deficiency, and interfering with VvDTX41B expression in grapes remarkably reduces the accumulation of proanthocyanidin. In addition, compared with the VvGSTU60-OE callus, the content of proanthocyanidin in VvDTX41B-RNAi + VvGSTU60-OE callus was significantly decreased but higher than that in VvDTX41B-RNAi callus. The results suggest that VvGSTU60 and VvDTX41B are coordinated in proanthocyanidin accumulation. These findings offer new insights into the accumulation mechanisms of proanthocyanidin in plants and provide the molecular basis for optimizing grape quality and wine production.