Differential regulation of xylem and phloem differentiation in grape berries by GA3 and CPPU

•GA3 and CPPU, either alone or in combination, caused an increase in berry size, area of berry vascular bundles, relative vessel number, and water transport in grape berry.•The CPPU treatment showed a preference for increasing phloem area of berry vascular bundles, whereas GA3 primarily increased the xylem area.•The GA3 treatment impacted specific transcription factors associated with xylem differentiation, whereas CPPU and the combined treatment triggered a diverse set of genes linked to the initiation, proliferation, and differentiation of vascular bundles. Fruit vascular bundles play a critical role in transporting water and nutrients within fruit, ultimately impacting fruit size and quality. Gibberellic acid (GA) and the synthetic cytokinin forchlorfenuron (CPPU) are commonly used to enhance grape berry size. Understanding their effects on the structure and function of berry vascular bundles could provide new information on the regulatory mechanism of these hormones on grapevine expansion and quality. In this study, the fruitlets of ‘Pinot Noir’ and ‘Einset Seedless’ grape varieties were treated by GA3, CPPU, and a combination of both at two weeks after full bloom. The samples were collected at 20, 40 and 70 days post-treatment. The results showed that all treatments led to an increase in berry size, xylem area, phloem area, the relative abundance of large-diameter grade vessels (>6 μm), the corresponding xylem area-specific hydraulic conductivity, and water absorption in both grape varieties. Specifically, CPPU favored an increase in phloem area, while GA3 primarily increased the xylem area. The combined application had a strong effect on both xylem and phloem area. RNA-seq analysis revealed that all treatments induced variations in the expression levels of multiple hormone signal components and genes related to vascular development. GA3 treatment mainly influenced some specific transcription factors involved in xylem differentiation, while CPPU and the combination treatment induced a wide range of genes related to vascular cambium initiation, proliferation, and differentiation of phloem and xylem vessels. Overall, our findings suggest that GA3 played a significant role in xylem differentiation, whereas CPPU had a major impact on both phloem and xylem differentiation in grape berries. These results lay the groundwork for further research the precise regulation of fruit vascular development under GA3 and CPPU treatments.