Cytological and molecular characterization of carotenoid accumulation in normal and high-lycopene mutant oranges

Ripe Cara Cara sweet orange contains 25 times as much carotenoids in flesh as Newhall sweet orange, due to high accumulation of carotenes, mainly phytoene, lycopene and phytofluene. Only yellow globular chromoplasts were observed in Newhall flesh. Distinct yellow globular and red elongated crystalline chromoplasts were found in Cara Cara but only one type of chromoplast was present in each cell. The red crystalline chromoplasts contained lycopene as a dominant carotenoid and were associated with characteristic carotenoid sequestering structures. The increased accumulation of linear carotenes in Cara Cara is not explained by differences in expression of all 18 carotenogenic genes or gene family members examined, or sequence or abundance of mRNAs from phytoene synthase (PSY) and chromoplast-specific lycopene β-cyclase (CYCB) alleles. 2-(4-Chlorophenylthio)-triethylamine hydrochloride (CPTA) enhanced lycopene accumulation and induced occurrence of red crystalline chromoplasts in cultured Newhall juice vesicles, indicating that carotenoid synthesis and accumulation can directly affect chromoplast differentiation and structure. Norflurazon (NFZ) treatment resulted in high accumulation of phytoene and phytofluene in both oranges, and the biosynthetic activity upstream of phytoene desaturase was similar in Newhall and Cara Cara. Possible mechanisms for high carotene accumulation and unique development of red crystalline chromoplasts in Cara Cara are discussed.