Natural overexpression of CAROTENOID CLEAVAGE DIOXYGENASE 4 in tomato alters carotenoid flux

초록

Carotenoids and apocarotenoids function as pigments and flavor volatiles in plants that enhance consumer appeal and offer health benefits. Tomato (Solanum lycopersicum.) fruit, especially those of wild species, exhibit a high degree of natural variation in carotenoid and apocarotenoid contents. Using positional cloning and an introgression line (IL) of Solanum habrochaites "LA1777', IL8A, we identified carotenoid cleavage dioxygenase 4 (CCD4) as the factor responsible for controlling the dark orange fruit color. CCD4b expression in ripe fruit of IL8A plants was similar to 8,000 times greater than that in the wild type, presumably due to 5 ' cis-regulatory changes. The ShCCD4b-GFP fusion protein localized in the plastid. Phytoene, zeta-carotene, and neurosporene levels increased in ShCCD4b-overexpressing ripe fruit, whereas trans-lycopene, beta-carotene, and lutein levels were reduced, suggestive of feedback regulation in the carotenoid pathway by an unknown apocarotenoid. Solid-phase microextraction-gas chromatography-mass spectrometry analysis showed increased levels of geranylacetone and beta-ionone in ShCCD4b-overexpressing ripe fruit coupled with a beta-cyclocitral deficiency. In carotenoid-accumulating Escherichia coli strains, ShCCD4b cleaved both zeta-carotene and beta-carotene at the C9-C10 (C9 '-C10 ') positions to produce geranylacetone and beta-ionone, respectively. Exogenous beta-cyclocitral decreased carotenoid synthesis in the ripening fruit of tomato and pepper (Capsicum annuum), suggesting feedback inhibition in the pathway. Our findings will be helpful for enhancing the aesthetic and nutritional value of tomato and for understanding the complex regulatory mechanisms of carotenoid and apocarotenoid biogenesis. CAROTENOID CLEAVAGE DIOXYGENASE 4b cleaves zeta-carotene and beta-carotene, and its overexpression leads to beta-cyclocitral deficiency in tomato fruit.

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