Functional Validation of a Phytoene Synthase and Lycopene ε-Cyclase Genes for High Lycopene Content in Autumn Olive Fruit (Elaeagnus umbellata).

Lycopene is the most potent antioxidant amongst all carotenoids and is beneficial for human health. The ripe fruit of autumn olive (Elaeagnus umbellata Thunb.) accumulate a high level of lycopene, which is 5- to 20-times higher than that of ordinary tomato. During the fruit ripening of autumn olive, only phytoene synthase (EutPSY) expression pattern shows a tight positive correlation with the increased lycopene content observed at four ripening stages, while the lycopene e-cyclase (EutLCYe) transcript could not be detected throughout fruit ripening. Here, we investigated whether the two genes are important targets for engineering lycopene biosynthesis. The full-length cDNAs of EutPSY and EutLCYe were firstly isolated. Fruit-specific overexpression of EutPSY in tomato fruits resulted in elevated contents of lycopene and β-carotene through feed-forward regulation of carotenogenic genes, i.e. down-regulation of SlLCYe and up-regulation of SlLCYb and SlCYCB. These fruit were decreased in ethylene production throughout ripening. Transcript levels of genes for system-2 ethylene synthesis (SlACS2, SlACS4, SlACO1 and SlACO3), perception (SlNR/ETR3 and SlETR4), and response (SlE4 and SlE8) were also inhibited in EutPSY-overexpressing fruit. Repressing ethylene synthesis and signaling transduction delayed fruit climacteric ripening of transgenic tomato plants. Additionally, RNAi suppression of SlLCYe enhanced β-carotene but not lycopene accumulation through altered expression of carotenogenic genes in transgenic tomato fruit by both feed-forward and feed-back regulatory mechanisms. Ethylene production in SlLCYe -RNAi fruit decreased, thereby delaying fruit ripening. Collectively, these results confirmed that transcriptional regulation of EutPSY and EutLCYe play a crucial role and a part in the massive lycopene accumulation in autumn olive fruit, respectively. EutPSY overexpression enhanced lycopene accumulation in tomato fruit independently of ethylene pathway, but did not influence the size and weight of tomato fruit. EutPSY can be used as an effective strategy capable of elevating lycopene content in fruit for improving quality.