BrPIF4/BrBBX24-BrHB52 mediated hypocotyl modularity unlocks mechanized harvesting potential in Brassica rapa

Chinese cabbage production faces critical mechanization challenges due to traditional plant architectures that limit mechanical harvesting efficiency. Traditional breeding prioritized short-hypocotyl varieties to prevent damping-off, but long hypocotyls are now critical for mechanical harvesting. We identified BrHB52, an HD-Zip transcription factor, as a key regulator of hypocotyl elongation through QTL mapping, RNA-seq, and haplotype analysis. BrHB52 expression was significantly higher in the long-hypocotyl variety R031L than in the short-hypocotyl variety R032S. Overexpression of BrHB52 in both Chinese cabbage and Arabidopsis led to elongated hypocotyls. The silencing of BrHB52 in R031L resulted in a reduction of hypocotyl length. Sequence alignment revealed a 251-bp insertion in the BrHB52 promoter of the long-hypocotyl variety R031L, which introduced the light-responsive GT-1 motifs. The upstream transcription factors Phytochrome-interacting factor4 (PIF4) and B-box zinc finger 24 (BBX24) were identified through yeast one-hybrid screening using the BrHB52R031L promoter sequence. PIF4 were found to bind to the both BrHB52R031L and BrHB52R032S promoters and activate their expression through G-box, while light-induced factor BBX24 only bind to the BrHB52R031L promoter and activate its expression by light-responsive element GT-1. Our findings elucidate a BrPIF4/BrBBX24-BrHB52 regulatory module that controls plant architecture through hypocotyl elongation. These findings not only provide critical genetic targets for developing mechanization-compatible Chinese cabbage, but also develop transgenic prototypes with elongated hypocotyls, offering practical resources for mechanized breeding.