Physiological metabolites and transcriptional regulation contribute to the feminization of mixed dormant buds at the tips of flowering mother branches in the persimmon cultivar ‘Taishuu’

• Phenylalanine metabolism pathway might have promoted persimmon female development. • Cytokinins likely promoted, while jasmonic acids inhibited persimmon feminization. • bHLH48 was suggested to be a key gene for the feminization in monoecious persimmon. Most persimmon ( Diospyros kaki Thunb.) cultivars are hexaploid, and the presence or absence of a Y chromosome strongly determines the plant's sexual type. Gynoecious plants, bearing only female flowers, lack a Y chromosome, whereas androecious (male-flowering) and monoecious (bearing both male and female flowers) plants possess a Y chromosome, creating a male-biased genetic profile. The Japanese pollination-constant, non-astringent persimmon cultivar ‘Taishuu’ is monoecious. In this cultivar, mixed dormant buds (MDBs) at the tips of robust flowering mother branches (FMBs) consistently develop into female shoots, whereas those at the basal portions of robust FMBs produce male shoots. The mechanism underlying the feminization of apical MDBs remains unclear. In this study, robust FMBs of ‘Taishuu’ were divided into four sections from apex to base, and a comparative metabolomic and transcriptomic analysis was conducted on MDBs from each section in mid-June. Additionally, levels of 109 phytohormones were measured across MDBs from the different sections. The results revealed that metabolites upregulated in MDBs at the tips of robust FMBs were significantly enriched in five metabolic pathways: arginine biosynthesis, beta-alanine metabolism, tropane/piperidine/pyridine alkaloid biosynthesis, ABC transporters, and phenylalanine metabolism. These pathways are associated with feminizing effects. Furthermore, cytokinins (CKs) and salicylic acids (SAs) generally promoted female development, whereas most jasmonic acids (JAs) appeared to inhibit female traits. Transcriptome analysis identified bHLH48 as a potential key regulator of feminization in MDBs at the tips of robust FMBs. This gene likely promotes feminization by increasing SAs, CKs, and flavonoid levels while concurrently reducing JAs. Overall, these findings provide new physiological and molecular insights into monoecious persimmon formation, which may support the artificial regulation of flower sexuality in ‘Taishuu’ and other monoecious persimmon cultivars.