<scp>BolAG</scp> ‐ <scp>BolRPI2</scp> ‐ <scp> <i>BolBAM4</i> </scp> Module Regulates the Bolting of Curd by Modulating Starch Metabolism in Broccoli ( <scp> <i>Brassica oleracea</i> </scp> var. <scp> <i>italica</i> </scp> )

The curd is a unique indeterminate inflorescence structure, which is the most important edible organ of broccoli. Premature bolting of the curd is a major factor that limits the yield and marketability of broccoli. However, the formation and development of broccoli curd, especially the bolting of curd, remain poorly understood. In this study, BolAG, a homologous gene of the floral development gene AtAG, was highly expressed in curd and floral organ of broccoli. Overexpression of BolAG in broccoli significantly promoted curd development, leading to early bolting. Correspondingly, RNA interference (RNAi) of BolAG resulted in arrested curd development, leading to delayed or even absent bolting. Physiological analyses indicated that BolAG-overexpressing broccoli exhibited higher starch and soluble sugar contents (SSC) compared to the wild-type (WT). RNA-Seq analysis also identified significant enrichment of genes related to starch metabolism. Among them, the starch hydrolysis-related gene BolBAM4 was confirmed to be the target gene of BolAG. Additionally, BolAG was confirmed to interact with ribose-5-phosphate isomerase 2 (BolRPI2), a protein involved in the pentose phosphate pathway and their interaction enhanced the transcriptional promotion of BolAG on BolBAM4, accelerating starch degradation and thereby leading to the accumulation of soluble sugars. While BolRPI2 was demonstrated to have a positive effect on chloroplast function and promote starch synthesis, BolAG appeared to promote starch synthesis in a BolRPI2-independent manner. These results confirm that the BolAG-BolRPI2-BolBAM4 module plays a crucial role in regulating starch metabolism, providing a theoretical foundation for addressing industrial losses due to premature bolting.