Anthocyanin accumulation and differential gene expression in wild-typeand mutant Syzygium malaccense fruits during their growth and ripening

Ripe wild-type Malay apple (Syzygium malaccense) cv. Saraek fruit epidermis is red, whereas a mutant has a white skin. Wild-type and mutant fruit were used as a fruit model to study the regulation and gene expression patterns of anthocyanin biosynthesis and a myeloblastosis (MYB) transcription factor during growth and development. Fruit size, color, anthocyanin composition, and the expression of genes involved in anthocyanin biosyntheses were determined during fruit growth and ripening. Wild-type ripe fruit had a greater diameter and fruit mass than the mutant. The total anthocyanin content was approximately nine-fold higher in wild-type red fruit than in mutant white fruit. The major anthocyanin in the fruit skin of the wild-type was cyanidin-3-O-glucoside with minor amounts of pelargonidin-3-O-glucoside, peonidin-3-O-glucoside, and cyanidin-3,5-O-diglucoside. No anthocyanin was found in mutant fruit. The accumulation of cyanidin-3-O-glucoside during fruit growth and ripening was correlated with red color development and activities of phenylalanine ammonia lyase (PAL) and UDP-glucose:flavonoid-3-O-glucosyltransferase (UFGT). We cloned fragments and characterized seven genes involved in anthocyanin biosynthesis pathway namely phenylalanine ammonia lyase (SmPAL), chalcone synthase (SmCHS), chalcone isomerase (SmCHI), flavanone-3-hydroxylase (SmF3H), dihydroflavonol 4-reductase (SmDFR), leucoanthocyanidin dioxygenase (SmLDOX), and UDP glucose-flavonoid-3-O-glucosyl transferase (SmUFGT), as well as a MYB transcription factor (SmMYB). The expressions of all the genes were determined by semi-quantitative reverse transcription (RT)-PCR and quantitative real-time RT-PCR. The skin of wild-type fruit contained transcripts of all above mentioned genes, whereas the mutant fruit showed no SmUFGT and SmMYB expressions, which correlated with the absence of anthocyanin accumulation during fruit growth and ripening. These results suggest that lack of anthocyanin biosynthesis in mutant fruit may be via the regulation of UFGT and MYB transcription factor expressions.