Abstract

Flavonoid biosynthesis is crucial for pigmentation and quality formation in ripening pineapple fruit; however, its transcriptional regulation remains poorly understood. Here, we identify AcBBX6 as a key positive regulator. We demonstrate that AcBBX6 activates the transcription of flavonoid pathway genes (AcCHI2, AcF3′5′H2, AcF3H, AcFLS2, AcPAL1, AcLAR, AcGST1) by binding to their G-box elements in the promoters. Furthermore, the interaction of AcBBX6 and AcHY5 synergistically enhances the activation of flavonoid biosynthesis. Intriguingly, AcHY5 also interacts with AcBBX4, forming a ternary AcBBX6-AcBBX4-AcHY5 complex with stronger activation than any individual component, but its activity did not exceed that of the binary complexes. Silencing of AcBBX6 or AcBBX4 led to reduced flavonoid accumulation, whereas overexpression enhanced it. Coordinated manipulation of all three factors produced the most pronounced phenotypic effects on flavonoid content. Collectively, our results define the AcBBXs-AcHY5 complex as a core transcriptional module governing flavonoid biosynthesis during pineapple fruit ripening.