Cloning of 9-cis-epoxycarotenoid dioxygenase (NCED) gene encoding a key enzyme during abscisic acid (ABA) biosynthesis and ABA-regulated ethylene production in detached young persimmon calyx

Unlike the typical climacteric fruits, persimmons (Diospyros kaki Thunb.) produce higher levels of ethylene when they are detached from trees at a younger stage. In order to obtain detailed information on the role of abscisic acid (ABA) in ripening, we cloned the DKNCED1, DKACS2, and DKACO1 genes from the calyx. Water loss was first noted in the calyx lobe, and DKNCED1 was highly expressed 1 d after the fruits were detached, coinciding with an increase in the ABA content. Then, the DKACS2 and DKACO1 genes were expressed after some delay. In the calyx, the ABA peak was observed 2 d after the fruits were harvested, and this peak preceded the ethylene peak observed on day 3. The fruit firmness rapidly decreased on day 4, and the fruits softened completely 6 d after they were harvested. The increases in the expressions of ABA, ethylene, and the genes in the calyxes occurred earlier than the corresponding increases in the pulp, although the 3 increases occurred on different days. Exogenous ABA treatment increased ABA concentration, induced expression of both ACS and ACO, and promoted ethylene synthesis and young-fruit softening; by contrast, treatment with NDGA inhibited the gene expressions and ethylene synthesis and delayed young-fruit softening. These results indicate that ethylene biosynthesis in the detached young persimmon fruits is initially triggered by ABA, which is induced by water loss in the calyx, through the induction of DKACS2 and DKACO1 expressions. The ethylene produced in the calyx subsequently diffuses into the pulp tissue, where it induces autocatalytic ethylene biosynthesis, resulting in an abrupt increase in ethylene production.